Complete canine spinal cord transection model: a large animal model for the translational research of spinal cord regeneration

正Traumatic spinal cord injury (SCI) usually results in devastating neurologic deficits and disability. In the United States,approximately 12,500 new cases are reported each year, while an estimated 100,000–140,000 new cases occur every year in China (National Spinal Cord Injury Statistical Center, 2016).Spinal cord injuries are highly disabling and primarily affect young adults, and therefore create great psychological and financial burden on the affected individuals and their families.     

The effect of hydrogel injection on cardiac function and myocardial mechanics in a computational post-infarction model

An emerging therapy to limit adverse heart remodelling following myocardial infarction (MI) is the injection of polymers into the infarcted left ventricle (LV). In the few numerical studies carried out in this field, the definition and distribution of the hydrogel in the infarcted myocardium were simplified. In this computational study, a more realistic biomaterial distribution was simulated after which the effect on cardiac function and mechanics was studied. A validated finite element heart model was used in which an antero-apical infarct was defined. Four infarct models were created representing different temporal phases in the progression of a MI. Hydrogel layers were simulated in the infarcted myocardium in each model. Biomechanical and functional improvement of the LV was found after hydrogel inclusion in the ischaemic models representing the early phases of MI. In contrast, only functional but no mechanical restitution was shown in the scar model due to hydrogel presence.     

Left ventricular twist is load-dependent as shown in a large animal model with controlled cardiac load

ABSTRACT: BACKGROUND: Left ventricular rotation and twist can be assessed noninvasively by speckle tracking echocardiography. We sought to characterize the effects of acute load change and change in inotropic state on rotation parameters as a measure of left ventricular (LV) contractility. METHODS: Seven anesthetised juvenile pigs were studied, using direct measurement of left ventricular pressure and volume and simultaneous transthoracic echocardiography. Transient inflation of an inferior vena cava balloon (IVCB) catheter produced controlled load reduction. First and last beats in the sequence of eight were analysed with speckle tracking (STE) during the load alteration and analysed for change in rotation/twist during controlled load alteration at same contractile status. Two pharmacological inotropic interventions were also included to examine the same hypothesis in additionally conditions of increased and decreased myocardial contractility in each animal. Paired comparisons were made for different load states using the Wilcoxon's Signed Rank test. RESULTS: The inferior vena cava balloon occlusion (IVCBO) load change compared for first to last beat resulted in LV twist increase (11.67degrees +/-2.65degrees vs. 16.17degrees +/-3.56degrees respectively, p < 0.004) during the load alteration and under adrenaline stimulation LV twist increase 12.56degrees +/-5.1degrees vs. 16.57degrees +/-4.6degrees (p < 0.013), and though increased, didn't reach significance in negative inotropic condition. Untwisting rate increased significantly at baseline from 41.7degrees/s +/-41.6degrees/s vs.122.6degrees/s +/-55.8degrees/s (P < 0.039) and under adrenaline stimulation untwisting rate increased (55.3degrees/s +/-3.8degrees/s vs.111.4degrees/s +/-24.0degrees/s (p < 0.05), but did not systematically changed in negative inotropic condition. CONCLUSIONS: Peak systolic LV twist and peak early diastolic untwisting rate are load dependent. Differences in LV load should be included in the interpretation when serial measures of twist are compared.     

Effects of milrinone on left ventricular cardiac function during cooling in an intact animal model

A significant reduction in immunogenicity has been observed in some frozen-thawed tissues after cryopreservation. The objective of this study was to evaluate the effects of programmed cryopreservation on immunogenicity of rabbit bladder mucosa and on the extent of immunological rejection caused by the allograft. This study would provide theoretical support for the application of allogenic frozen-thawed bladder mucosa in the treatment of urethral stricture. Forty-two adult male New Zealand rabbits were used in this study. The immunogenicity was detected by mixed lymphocyte reaction using the allograft of bladder mucosa (fresh and frozen-thawed) and spleen lymphocytes. Twelve urethral stricture models were established in New Zealand rabbits for substitution urethroplasty using the allograft of bladder mucosa, which were divided into fresh and frozen-thawed group. Two weeks after operation, lymphocyte proliferation was detected in both blood and spleen of recipient rabbits. At the same time, immunohistochemical staining of urethral allograft was performed and the expression of CD3, CD4 and CD8 were observed. The mRNA of bladder mucosa (fresh and frozen-thawed) was extracted and the expressions of RLA-I, RLA-II and RLA-III gene were detected by real-time PCR. By mixed lymphocyte reaction, we found that allogenic lymphocyte proliferation stimulated by frozen-thawed bladder epithelial cells was significantly weaker than that of the fresh cells. The blood and spleen lymphocytes from fresh bladder mucosa group showed significantly higher proliferation rate than frozen-thawed group. Compared with the fresh group, the expression of CD3+ and CD8+ T cells infiltrated in the operation locus of bladder mucosa urethroplasty was significantly decreased in the frozen-thawed group. However, the expressions of RLA genes did not change significantly after the freeze-thaw procedure. This study demonstrates for the first time that a programmed freeze-thaw procedure of rabbit bladder mucosa could reduce its immunogenicity in allogenic bladder mucosa urethroplasty and thus restrict the extent of immunological rejection, therefore, provides theoretical support for the application of frozen-thawed bladder mucosa in the treatment of urethral stricture.     

The cardiac mutant Mexican axolotl is a unique animal model for evaluation of cardiac myofibrillogenesis

Hearts from cardiac mutant Mexican axolotl, Ambystoma mexicanum, do not form organized myofibrils and fail to beat. Though previous biochemical and immunohistochemical experiments showed a possible reduction of cardiac tropomyosin it was not clear that this caused the lack of organized myofibrils in mutant hearts. We used cationic liposomes to introduce both rabbit and chicken tropomyosin protein into whole hearts of embryonic axolotls in whole heart organ cultures. The mutant hearts had a striking increase in the number of well-organized sarcomeric myofibrils when treated with rabbit or chicken tropomyosin. FITC-labeled rabbit tropomyosin was used to examine the kinetics of incorporation of the exogenous protein into mutant hearts and confirmed the uptake of exogenous protein by the cells of live hearts in culture. By 4 h of transfection, both normal and mutant hearts were found to incorporate FITC-labeled tropomyosin into myofibrils. We also delivered an anti-tropomyosin antibody (CH 1) into normal hearts to disrupt the existing cardiac myofibrils which also resulted in reduced heartbeat rates. CH1 antibody was detected within the hearts and disorganization of the myofibrils was apparent when compared to normal controls. Introduction of a C-protein monoclonal antibody (ALD 66) did not result in a disruption of organized myofibrils. The results show clearly that chicken or rabbit tropomyosin could be incorporated by the mutant hearts and that it was sufficient to overcome the factors causing a lack of myofibril formation in the mutant. This finding also suggests that a lack of organized myofibrils is caused primarily by either inadequate levels of tropomyosin or endogenous tropomyosin in mutant hearts is unsuitable for myofibril formation, which we were able to duplicate with the introduction of tropomyosin antibody. Furthermore, incorporation of a specific exogenous protein or antibody into normal and mutant hearts of the Mexican axolotl in whole heart organ culture offers an unique model to evaluate functionalroles of contractile proteins necessary for cardiac development and differentiation.     

Impact of anesthesia on cardiac function during echocardiography in mice

Anesthetics provide sedation and immobility facilitating echocardiography in mice, but influence cardiac function. We studied the effects of intraperitoneal and inhaled anesthetic agents on echocardiographic measurements. Mice were anesthetized with intraperitoneal tribromoethanol (TBE), ketamine-midazolam (K/M), ketamine-xylazine (K/X), or inhaled isoflurane (Isf), and echocardiographic parameters were assessed at 5, 10, 15, and 20 min. In C57BL/6N mice, Isf produced high initial heart rates (HR) that decreased to levels comparable to TBE at 15-20 min (approximately 450 beats/min) and the most stable percent fractional shortening (%FS) and end-diastolic dimension (EDD). With TBE, %FS initially was low, but increased comparable to Isf (approximately 45%) at 15 min. K/M produced similar time trends but lower absolute values compared with TBE for all parameters. K/X produced cardiac depression evidenced by low HR and %FS, and increased EDD. Isf was the most reproducible in repeat studies at 12 days. In C57BL/6J compared with C57BL/6N mice, K/M produced higher HR, and %FS and TBE produced smaller EDD. In conclusion, anesthetic agent, timing of echocardiographic measurements, and genetic background are all critical variables during echocardiography in mice.     

Comparative analysis of parvalbumin and SERCA2a cardiac myocyte gene transfer in a large animal model of diastolic dysfunction

Diastolic dysfunction results from impaired ventricular relaxation and is an important component of human heart failure. Genetic modification of intracellular calcium-handling proteins may hold promise to redress diastolic dysfunction; however, it is unclear whether other important aspects of myocyte function would be compromised by this approach. Accordingly, a large animal model of humanlike diastolic dysfunction was established through 1 yr of left ventricular (LV) pressure overload by descending thoracic aortic coarctation in canines. Serial echocardiography documented a progressive increase in LV mass. Diastolic dysfunction with preserved systolic function was evident at the whole organ and myocyte levels in this model, as determined by hemispheric sonomicrometric piezoelectric crystals, pressure transducer catheterization, and isolated myocyte studies. Gene transfer of the sarco(endo)plasmic reticulum calcium-ATPase (SERCA2a) and parvalbumin (Parv), a fast-twitch skeletal muscle Ca(2+) buffer, restored cardiac myocyte relaxation in a dose-dependent manner under baseline conditions. At high Parv concentrations, sarcomere shortening was depressed. In contrast, during beta-adrenergic stimulation, the expected enhancement of myocyte contraction (inotropy) was abrogated by SERCA2a but not by Parv. The mechanism of this effect is unknown, but it could relate to the uncoupling of SERCA2a/phospholamban in SERCA2a myocytes. Considering that inotropy is vital to overall cardiac performance, the divergent effects of SERCA2a and Parv reported here could impact potential therapeutic strategies for human heart failure.     

糖尿病动物模型心功能评价方法

糖尿病心血管并发症（ cardiovascular complications of diabetes , CCD）是糖尿病患者最主要的死亡原因,其中糖尿病心肌病（ diabetic cardiomyopathy , DC）是心力衰竭的主要原因。对于分析糖尿病心肌病的机制、早期诊断以及改进和优化治疗,心脏功能评估起到重要桥梁作用,而糖尿病动物模型直接或间接反映糖尿病的发生和发展过程,是一个良好的研究载体。本文旨在综述国内外糖尿病动物模型心脏功能评价的重要方法。     

腰麻-硬膜外联合麻醉及全凭静脉用于经皮肾镜碎石术的麻醉效果比较

目的观察经皮肾镜碎石术行腰麻-硬膜外联合麻醉与全凭静脉麻醉的临床效果。方法选择我院行经皮肾镜碎石术治疗的80例患者为研究以对象,随机分为对照组和研究组各40例,对照组实施全凭静脉麻醉,研究组实施腰麻-硬膜外联合麻醉,观察比较两组患者的麻醉效果、麻醉完全苏醒时间和不良反应情况。结果研究组的麻醉总有效率达95.0%明显高于对照组的77.5%,不良反应少于对照组,麻醉完全苏醒时间短于对照组,差异均有统计学意义(均P0.05)。结论经皮肾镜碎石术采用腰麻-硬膜外联合麻醉的效果好,苏醒时间短,不良反应少,是一种安全可行的麻醉方式。     

Effects of avertin versus xylazine-ketamine anesthesia on cardiac function in normal mice

Anesthetic regimens commonly administered during studies that assess cardiac structure and function in mice are xylazine-ketamine (XK) and avertin (AV). While it is known that XK anesthesia produces more bradycardia in the mouse, the effects of XK and AV on cardiac function have not been compared. We anesthetized normal adult male Swiss Webster mice with XK or AV. Transthoracic echocardiography and closed-chest cardiac catheterization were performed to assess heart rate (HR), left ventricular (LV) dimensions at end diastole and end systole (LVDd and LVDs, respectively), fractional shortening (FS), LV end-diastolic pressure (LVEDP), the time constant of isovolumic relaxation (tau), and the first derivatives of LV pressure rise and fall (dP/dt(max) and dP/dt(min), respectively). During echocardiography, HR was lower in XK than AV mice (250 +/- 14 beats/min in XK vs. 453 +/- 24 beats/min in AV, P < 0.05). Preload was increased in XK mice (LVDd: 4.1 +/- 0.08 mm in XK vs. 3.8 +/- 0.09 mm in AV, P < 0.05). FS, a load-dependent index of systolic function, was increased in XK mice (45 +/- 1.2% in XK vs. 40 +/- 0.8% in AV, P < 0.05). At LV catheterization, the difference in HR with AV (453 +/- 24 beats/min) and XK (342 +/- 30 beats/min, P < 0.05) anesthesia was more variable, and no significant differences in systolic or diastolic function were seen in the group as a whole. However, in XK mice with HR <300 beats/min, LVEDP was increased (28 +/- 5 vs. 6.2 +/- 2 mmHg in mice with HR >300 beats/min, P < 0.05), whereas systolic (LV dP/dt(max): 4,402 +/- 798 vs. 8,250 +/- 415 mmHg/s in mice with HR >300 beats/min, P < 0.05) and diastolic (tau: 23 +/- 2 vs. 14 +/- 1 ms in mice with HR >300 beats/min, P < 0.05) function were impaired. Compared with AV, XK produces profound bradycardia with effects on loading conditions and ventricular function. The disparate findings at echocardiography and LV catheterization underscore the importance of comprehensive assessment of LV function in the mouse.     

Repolarization abnormalities and afterdepolarizations in a canine model of sudden cardiac death

Ventricular tachyarrhythmias are the most common cause of sudden cardiac death (SCD); a healed myocardial infarction increases the risk of SCD. We determined the contribution of specific repolarization abnormalities to ventricular tachyarrhythmias in a postinfarction model of SCD. For our methods, we used a postinfarction canine model of SCD, where an exercise and ischemia test was used to stratify animals as either susceptible (VF(+)) or resistant (VF(-)) to sustained ventricular tachyarrhythmias. Our results show no changes in global left ventricular contractility or volumes occurred after infarction. At 8-10 wk postmyocardial infarction, myocytes were isolated from the left ventricular midmyocardial wall and studied. In the VF(+) animals, myocyte action potential (AP) prolongation occurred at 50 and 90% repolarization (P < 0.05) and was associated with increased variability of AP duration and afterdepolarizations. Multiple repolarizing K(+) currents (I(Kr), I(to)) and inward I(K1) were also reduced (P < 0.05) in myocytes from VF(+) animals compared with control, noninfarcted dogs. In contrast, only I(to) was reduced in VF(-) myocytes compared with controls (P < 0.05). While afterdepolarizations were not elicited at baseline in myocytes from VF(-) animals, afterdepolarizations were consistently elicited after the addition of an I(Kr) blocker. In conclusion, the loss of repolarization reserve via reductions in multiple repolarizing currents in the VF(+) myocytes leads to AP prolongation, repolarization instability, and afterdepolarizations in myocytes from animals susceptible to SCD. These abnormalities may provide a substrate for initiation of postmyocardial infarction ventricular tachyarrhythmias.     

Partial gravity unloading inhibits bone healing responses in a large animal model

The reduction in mechanical loading associated with space travel results in dramatic decreases in the bone mineral density (BMD) and mechanical strength of skeletal tissue resulting in increased fracture risk during spaceflight missions. Previous rodent studies have highlighted distinct bone healing differences in animals in gravitational environments versus those during spaceflight. While these data have demonstrated that microgravity has deleterious effects on fracture healing, the direct translation of these results to human skeletal repair remains problematic due to substantial differences between rodent and human bone. Thus, the objective of this study was to investigate the effects of partial gravitational unloading on long-bone fracture healing in a previously-developed large animal Haversian bone model. In vivo measurements demonstrated significantly higher orthopedic plate strains (i.e. load burden) in the Partial Unloading (PU) Group as compared to the Full Loading (FL) Group following the 28-day healing period due to inhibited healing in the reduced loading environment. DEXA BMD in the metatarsus of the PU Group decreased 17.6% (p<0.01) at the time of the ostectomy surgery. Four-point bending stiffness of the PU Group was 4.4 times lower than that of the FL Group (p<0.01), while µCT and histomorphometry demonstrated reduced periosteal callus area (p<0.05), mineralizing surface (p<0.05), mineral apposition rate (p<0.001), bone formation rate (p<0.001), and periosteal/endosteal osteoblast numbers (p<0.001/p<0.01, respectively) as well as increased periosteal osteoclast number (p<0.05). These data provide strong evidence that the mechanical environment dramatically affects the fracture healing cascade, and likely has a negative impact on Haversian system healing during spaceflight.     

Interaction frequency as a surrogate for the total effect of animal mutualists on plants

We evaluate whether species interaction frequency can be used as a surrogate for the total effect of a species on another. Because interaction frequency is easier to estimate than per‐interaction effect, using interaction frequency as a surrogate of total effect could facilitate the large‐scale analysis of quantitative patterns of species‐rich interaction networks. We show mathematically that the correlation between interaction frequency (I) and total effect (T) becomes more strongly positive the greater the variation of I relative to the variation of per‐interaction effect (P) and the greater the correlation between I and P. A meta‐analysis using data on I, P and T for animal pollinators and seed dispersers visiting plants shows a generally strong, positive relationship between T and I, in spite of no general relationship between P and I. Thus, frequent animal mutualists usually contribute the most to plant reproduction, regardless of their effectiveness on a per‐interaction basis.     

Levels of membrane fluidity in the spinal cord and the brain in an animal model of amyotrophic lateral sclerosis

A mutant form of the copper/zinc superoxide dismutase (SOD1) protein is found in some patients with amyotrophic lateral sclerosis (ALS). Alteration of the activity of this antioxidant enzyme leads to an oxidative stress imbalance, which damages the structure of lipids and proteins in the CNS. Using fluorescence spectroscopy, we monitored membrane fluidity in the spinal cord and the brain in a widely used animal model of ALS, the SOD^G93A mouse, which develops symptoms similar to ALS with an accelerated course. Our results show that the membrane fluidity of the spinal cord in this animal model significantly decreased in symptomatic animals compared with age-matched littermate controls. To the best of our knowledge, this is the first report showing that membrane fluidity is affected in the spinal cord of a SOD^G93A animal model of ALS. Changes in membrane fluidity likely contribute substantially to alterations in cell membrane functions in the nervous tissue from SOD^G93A mice.     

The effect of a high-intensity radiation exposure on the brain function of monkeys. The postradiation changes in brain bioelectrical activity

In experiments with Macaca fascicularis it has been found that changes in the total bioelectric activity of the brain within the EEG range that occur during the first 60 min following whole-body irradiation with a dose of 45 Gy (6.5 Gy/s) are interrelated with the dynamics of nervous and psychic activity of the exposed animals, exhibit a definite stereotype of their development (disorganization of rhythms: generalized synchronization of biopotentials; and development of synchronous processes), and coincide in time with the main stages of the development of the clinical picture of the acute postirradiation period (noncoordinated stimulation, sopor or coma, and partial recovery).     

Pyramidal tract function during onset of brain death

In 51 patients with primary brain lesions, who fulfilled the criteria of brain death, sequential recording of transcranial magnetic evoked potentials (TMEPs) and somatosensory evoked potentials (SEPs) were performed.In all comatose patients with apnoeic cranial nerve areflexia the TMEP could not be elicited, while the response after cervical magnetic stimulation was always preserved. Similarly, no cortical SEPs were preserved in apnoeic cranial nerve areflexia; however, the cervical somatosensory response was preserved in 44%.In deteriorating patients with coma grade III TMEPs were preserved in 3 instances, while cortical SEPs were already absent. Current brain death criteria, however, were not challenged, as TMEPs were absent in all 51 patients, at the latest when apnoea was noted.     

Biodistribution and clearance of intra-articular liposomes in a large animal model using a radiographic marker

The intra-articular (IA) route of administration in treating arthritis has potential for targeting drug delivery to affected tissues, thereby minimising the attendant side-effects of systemically administered drugs. The ultra-structure of the synovium however facilitates rapid drug efflux from the joint; effectively the IA route is equivalent to other non-IV parenteral routes with regards absorption and redistribution into the systemic circulation. The aim of this study was to extend the drug residence time within the knee joint by using a liposome formulation. DPPC-based liposomes were prepared with the radio contrast agent iohexol as a drug marker. 8 sheep had their right knees injected IA with iohexol liposomes and the contralateral joints with either free iohexol or empty liposomes. Joints were radiographed at multiple time points up to 16 days post-injection. Iohexol-mediated radiopacity was quantified by densitometer. Sheep were sacrificed at the end of the study for microscopy of synovial tissues. Good visualization of iohexol-mediated radiopacity with fine anatomical definition was possible throughout the experiment. Also evident on the films was extra-articular radiopacity with liposomes tracking along muscle facial planes. Cellular and tissue localization with light microscopy was possible through use of frozen sections and because of the large liposome size. Residence of encapsulated iohexol within the knee joint was greatly prolonged. Liposomal iohexol declined bi-exponentially with a terminal elimination half-life of 134 hours. In contrast, free iohexol was undetectable at 3 hours post-injection.     

Effect of flecainide on atrial fibrillatory rate in a large animal model with induced atrial fibrillation

Background

Atrial fibrillatory cycle length has been considered one of the indices of atrial electrical remodelling during atrial fibrillation (AF), which can be assessed from surface ECG by computer-assisted calculation of atrial fibrillatory rate (AFR). Horses have been suggested as a bona fide model for AF studies since horses too, develop lone AF, however data on AF characteristics in horses are extremely sparse and non-invasive characterization of AF complexity using surface ECG processing has not been reported.

Aim

The aim was to study characteristics of induced AF and its modification by flecainide.

Methods

The study group consisted on 3 horses with spontaneous persistent AF and 13 with pace-induced AF. Seven horses were treated with saline (control) and eight with flecainide (2 mg/kg). ECGs were analysed using spatiotemporal cancellation of QRST complexes and calculation of AFR from the residual atrial signal.

Results

At AF onset, AFR was 295?±?52 fibrillations per minute (fpm) in the horses with induced AF treated with flecainide, 269?±?36 fpm in the control group (ns), and 364?±?26 fpm in the horses with spontaneous persistent AF (P?<?0.05 compared to the control group). Flecainide caused a decrease in AFR in all animals and restored sinus rhythm in the animals with induced AF. In the control animals, AFR increased from 269?±?36 fpm to a plateau of 313?±?14 fpm before decreasing to 288?±?28 fpm during the last 10% of the AF episodes preceding spontaneous conversion (P?<?0.05).

Conclusion

AFR in horses with induced AF resembles AFR in humans with paroxysmal AF. Flecainide caused a rapid decrease in AFR in all horses, further supporting the method to be a non-invasive technique to study the effect of antiarrhythmic compounds.