The authors would like to thank Dr., Robert Bloch for his generous donation of laboratory space and facilities and Dr. Rao Gullapalli and Da Shi in the Core for Translational Imaging at Maryland (C-TRIM) and the Magnetic Resonance Research Center (MRRC) for technical support. This work was supported by grants to RML from the National Institutes of Health (K01AR053235 and 1R01AR059179) and from the Muscular Dystrophy Association (#4278), and by a grant to JAR from the Jain Foundation.

1. in vivo injury Model and measurement of isometric torque.
<ol>
 <li> These procedures can be used for rats or mice7,17,18. To begin, place the animal supine under inhalation anesthesia (~ 4-5% isoflurane 
 for induction in an induction chamber,then ~ 2% isoflurane via a nosecone for maintenance) using a precision vaporizer (cat # 91103, Vet Equip, Inc, 
 Pleasanton, CA). Apply sterile ophthalmic cream (Paralube Vet Ointment, PharmaDerm, Floham Park, NJ) to each eye to protect the corneas from drying. 
 During the procedure, the animal is kept warm by use of a heat lamp placed outside the cage and kept at least 6 inches from the animal at all times. </li>
 <li> Prep the skin by removing hair and by cleaning with alternating scrubs of betadine and 70% alcohol to prevent seeding skin bacteria into the soft tissue or bone. Confirm proper anesthesia by lack of a deep tendon reflex (no foot withdrawal in response to pinching the foot). A needle is manually placed through the proximal tibia in order to stabilize the limb onto the rig (25G or 27G for mouse). The needle should not enter the anterior compartment of the leg. </li>
 <li> Lock the needle into a fixed position, such that the animal is supine and the toes are facing straight up. A custom-made device is used to secure the needle and thereby stabilize the leg. </li>
 <li> Place the foot of the limb onto a custom-machined footplate (Figure 1). The axis of the footplate is attached to a stepper motor (model T8904, NMB Technologies, Chatsworth, CA) and a torque sensor (model QWFK-8M, Sensotec, Columbus, OH). The foot should initially be aligned so that it is orthogonal to the tibia, as in Figure 1. </li>
 <li> Use transcutaneous electrodes (723742, Harvard Apparatus, Cambridge, MA) or subcutaneous electrodes (J05 Needle Electrode Needles, 36BTP, Jari Electrode 
 Supply, Gilroy, CA) to stimulate the fibular nerve near the neck of the fibula, where the nerve lies in a superficial position. Visually confirm isolated 
 dorsiflexion by performing a series of twitches (0.1 ms pulse for the mouse and 1 ms pulse for the rat) before the foot is secured. Once the foot is secured 
 to the footplate with adhesive tape, an increase in twitch amplitude in response to an increase in voltage confirms that opposing muscles (plantarflexors) are not 
 being simultaneously stimulated. </li>
 <li> Before injury, and at selected time points after an injury, the maximal force producing capacity of the dorsiflexors is recorded as the 
 "maximal isometric torque" (torque without a change in muscle length). Torque measurements are performed on the same rig that is used to induce injury. Before recording maximal isometric torque, the pulse amplitude is adjusted to optimize twitch tension and the optimal position of the ankle is determined by giving twitches at different lengths of the dorsiflexors. After obtaining a torque-angle curve to determine the optimal length of the dorsiflexors (resting length, aka Lo), a torque frequency plot is obtained by progressively increasing the frequency of pulses during a 200 ms pulse train. A maximal fused tetanic contraction is obtained usually at 90-100 Hz. Three separate twitches and tetanic contractions are recorded and saved for further analysis. </li>
 <li> Use commercial software to (Labview version 8.5, National Instruments, Austin, TX) to synchronize contractile activation, onset of ankle rotation, and torque data collection. Stimulation of the dorsiflexor muscles occurs while the computer-controlled motor simultaneously moves the footplate into plantar flexion, thus leading to a lengthening contraction (also called "eccentric" contraction, which causes injury of the muscle). The specific protocol depends on the desired magnitude of injury desired by the investigator. The magnitude of injury, or tissue damage, can be regulated by manipulation of variables such as angular velocity, timing of muscle activation, range of motion, and the number of lengthening contractions. </li>
 <li> To induce injury, superimpose a lengthening contraction onto a maximal isometric contraction, varying the range of motion, velocity of lengthening, and timing of stimulation as needed. For example, a maximal isometric contraction is obtained in the dorsiflexors and after 200 ms they are lengthened at a selected velocity to approximate normal movement (900&deg;/sec). We have previously shown that activation before the movement and the degree of lengthening are important factors in obtaining an injury14. The majority of torque produced by the dorsiflexors is from the TA11 and we have shown previously that this model results in injury to this muscle5,13-15. The TA remains stimulated throughout lengthening. </li>
 <li> After injury, the animal is removed from the apparatus. The tibial pin is removed, the leg is cleaned again, and the animal is returned to the cage 
 (placed on a temperature-controlled heating block at 37&deg;C) and monitored until recovery. This includes waiting until the animal is awake and mobile. The animals suffer no observable pain during the procedure and there are no visible changes in gait (e.g. lameness) after injury induced by lengthening contractions. However, appropriate anti-pain treatment is administered subsequently (buprenorphine 0.05 - 0.1 mg/kg every 12 hours for 48 hr after surgery). </li>
</ol>
2. In situ measurement of whole muscle tension.
<ol>
 <li> The animal is prepared and the tibia is stabilized as described above in section 1.1 through 1.3. All instrumentation is turned on at least 30 min prior to testing for proper calibration and to minimize thermal drift of the force transducer. </li>
 <li> Incise the skin anterior to the ankle and sever the tendon of the tibialis anterior muscle (TA). Carefully tie 4.0 Ethicon silk non-absorbable suture to the tendon and attach the vicryl suture to the load cell via the provided S-hook (weight = 0.1g), model FT03, Grass Instruments, Warwick, RI). Alternatively, a custom clamp (weight = 0.5g) can be used to attach the tendon to the vicryl suture (Figure 2). </li>
 <li> The load cell is mounted to a micromanipulator (Kite Manipulator, World Precision Instruments Inc, Sarasota, FL) so that the TA could be adjusted to resting length and aligned properly (a straight line of pull between the origin and insertion). The TA is protected from cooling by a heat lamp and from dehydration by mineral oil. The signals from the load cell (calibrated before each test) are fed via a DC amplifier (model P122, Grass Instruments, Warwick, RI) to an A/D board to be collected and stored by acquisition software (PolyVIEW version 2.1, Grass Instruments, Warwick, RI). </li>
 <li> Attach the TA to the load cell and apply single twitches (rectangular pulse of 1 ms) at different muscle lengths in order to determine L0. Muscle resting length, measured using calipers, is defined as the distance between the tibial tuberosity and the myotendinous junction. At this length, gradually increase the pulse amplitude and then the pulse frequency to establish a force-frequency relationship. A maximally fused tetanic contraction is obtained at approximately 90-100Hz (300 ms train duration comprised of 0.1 ms or 1 ms pulses). Use 150% of the maximum stimulation intensity to activate the TA in order to induce maximal contractile activation (P0). Maximal tetanic contractions can be performed repeatedly and expressed as percentage of P0, providing an index of fatigue at any desired point in time. </li>
</ol>
3. in vivo MR imaging and/or spectroscopy of rodent skeletal muscles.
 All MRI and MRS is performed on a Bruker Biospin (Billerica, MA) 7.0 Tesla MR system equipped with a 12 cm gradient insert (660 mT/m maximum gradient, 4570 T/m/s maximum slew rate) running Paravision 5.0 software.
<ol>
 <li> The animal is anesthetized with vaporized isoflurane as described above in 1.1. An MR-compatible small-animal monitoring and gating system (SA Instruments, Inc.) is used to monitor respiration rate and body temperature. Mouse body temperature is maintained at 36-37&deg;C using a warm water circulator. A custom-made holder is used to position the mouse in the supine position with both legs parallel to the bore of the magnet from knee to foot. A four-channel receive-only surface coil is placed within a 72 mm linear 1H resonator. The resonator coil is tuned and matched to the sample. </li>
 <li> MR Imaging: After localizers, the following MR scans are performed: T1-weighted rapid acquisition with relaxation enhancement (RARE) with the following parameters: TE = 9.52 ms, TR = 1800, echo train length = 4, in-plane resolution 100x100 &mu;m, and slice thickness = 750 &mu;m. Dual-echo PD/T2 RARE: TE=19.0/57.1 ms, TR=5000 ms, echo train length = 4, in-plane resolution 100x100 &mu;m, and slice thickness = 750 &mu;m. Spin echo (SE) diffusion tensor image data using 12 non-colinear directions: b-value = 350 s/mm&#x2013;2, TE = 26 ms, TR = 4500 ms, in-plane resolution 150x150 &mu;m, and slice thickness = 750 &mu;m. Multi-slice multi-echo (MSME) T2 parametric mapping data using 16 TEs = 11.4 ms to 182.5 ms with &Delta;TE = 11.4 ms, TR = 10000 ms, in-plane resolution 150x150 &mu;m, and slice thickness = 750 &mu;m. </li>
 <li> Image processing: Diffusion tensor reconstruction and tractography is performed using TrackVis (Martinos Center for Biomedical Imaging; Massachusetts General Hospital; Boston, MA) to create mean diffusivity (MD), fractional anisotropy (FA) images as well as tractography maps. T2 mapping is performed using custom software written in MATLAB (The Mathworks; Natick, MA) using non-linear least squares to fit the measured data at each pixel to the canonical T2 signal equation. Regions of interest measurements are performed to assess parameter values within the TA. </li>
 <li> 1H spectroscopy: Automated shimming is performed on a 1 x 1 x 4 mm3 voxel in the TA. A point-resolved spectroscopy (PRESS) pulse sequence (TR/TE=2000/18 ms) is used to acquire spectra from the same voxel with 1024 averages. Data acquisition is 34 minutes on each leg. Spectral data are processed using the LCModel package16. 31P spectroscopy: A dual-tuned (1H, 31P) surface coil is used to perform non-localized (using a single-pulse experiment) or localized spectroscopy using the image selected in vivo spectroscopy (ISIS) pulse sequence. </li>
</ol>
4. Harvesting and storing muscles.
 TAs are harvested after at the end of experiments, weighed, snap frozen in liquid nitrogen, and then stored at -80&deg;C. This can be performed at any point in time after the in vivo experiments. Muscles are harvested immediately after the in situ experiments, as this is a terminal procedure. For detailed morphological studies, the animal is fixed with 4% paraformaldehyde via perfusion through the left ventricle. 
5. Representative results.
 Figure 3 shows representative data from a rat in the in vivo apparatus. The in vivo apparatus is used to obtain maximal torque generated by the dorsiflexor muscles; it is also used to induce injury to these same muscles. Due to the length-tension relationship of muscle, maximal isometric torque typically occurs when the ankle joint is positioned at approximately 20&deg; of plantarflexion (with the foot positioned orthogonal to the tibia considered 0&deg;). After maximal isometric torque is obtained, the foot can then be placed into any position to begin the injury protocol. Figure 3 represents an injury protocol of 30 repetitions with an arc of motion from 0&deg; - 70&deg;. Note the steady decline in torque generated from the isometric phase (filled arrow) and lengthening phase (open arrow) during the contraction-induced injury protocol. Torque is recorded in units of Nmm, but the absolute value depends on the size of the animal and its condition (e.g., injured muscle, fatigued muscle, or muscle lacking a certain protein due to homologous recombination). 
 Figure 4 shows representative data from a rat in the in situ apparatus. Our in situ apparatus does not involve lengthening contractions; rather, it allows us to isolate, properly align, and measure maximal tension produced by an individual muscle at a known length. Figure 4 shows the gradual loss of force that occurs during a fatigue test in a tibialis anterior muscle of a rat. In this particular example, titanic contractions were performed once every second for 5 minutes. Tension (force) is typically recorded in Newtons (or grams), but like torque, the absolute value depends on the size and condition of the animal. Because muscle weight is obtained immediately after this procedure, the force can be normalized (called "specific force") to muscle cross sectional area. 
 Figure 5 shows representative data from in vivo imaging of a mouse, such as T1-weighted and T2 parametric mapping (A), 3D tractography from diffusion tensor imaging (B), 1H spectroscopy (C), and 31P spectroscopy. Details are provided in the figure legend. 
 <img src="/files/ftp_upload/2782/2782fig1.jpg" alt="Figure 1" /> 
 Figure 1: in vivo apparatus.* To produce the injury, the tibia is stabilized and the foot attached to a motor-driven plate. The ankle dorsiflexors are stimulated via the fibular nerve while the footplate forces the foot into plantar flexion (dotted arrow). 
 * Lovering &amp; De Deyne, J Biomechanics 2005, used with permission. 
 <img src="/files/ftp_upload/2782/2782fig2.jpg" alt="Figure 2" /> 
 Figure 2: In situ apparatus The load cell is mounted to a micromanipulator so that the TA could be adjusted to resting length and aligned properly in the X, Y, and Z directions. The distal tendon of the TA is attached to the load cell and single twitches are induced at different muscle lengths in order to determine L0. A maximal tetanic contraction is obtained to determine maximal contractile activation (P0). Maximal tetanic tension can be performed repeatedly and expressed as percentage of P0, providing an index of fatigue at a desired point in time. 
 <img src="/files/ftp_upload/2782/2782fig3.jpg" alt="Figure 3" /> 
 Figure 3: Torque data from in vivo apparatus Representative trace recordings of torque from lengthening contractions in the rat. In this particular example, muscles were stimulated for 200 milliseconds to induce a peak isometric contraction (filled arrow) before lengthening (open arrow) by the footplate through a 70&deg; arc of motion at an angular velocity of 900&deg;/s. 
 <img src="/files/ftp_upload/2782/2782fig4.jpg" alt="Figure 4" /> 
 Figure 4: Tension data from in situ apparatus Representative data showing the decline in maximal isometric tetanic tension during repeated stimulation of the tibialis anterior muscle (TA) in a rat. In this example, the TA was isolated, adjusted to optimal length (L0), and then stimulated with a 200 ms tetanic contraction once every second for 5 minutes. 
 <img src="/files/ftp_upload/2782/2782fig5.jpg" alt="Figure 5" /> 
 Figure 5: in vivo imaging A: The images show transverse (axial) sections of T1-weighted and T2 parametric mapping from the tibialis anterior muscle (TA). The dotted red box surrounds the TA to show increased increased T2 in the injured (left side) versus uninjured (right side). B: Representative 3D tractography from diffusion tensor imaging (DTI). C: The 1H spectrum of a mouse TA shows several detectable lipid resonances; differentiation between intramyocellular (IMCL) and extramyocellular lipid (EMCL) peaks is obtained using this method. D: The 31P MR spectrum of the rat TA shows phosphocreatine (PCr), inorganic phosphate (Pi), and the three resonances (&alpha;, &beta;, &gamma;) of adenosine 5'-triphosphate (ATP).

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No conflicts of interest declared.

"Muscle damage" has been defined and measured in many ways. Structural damage is evident in histological findings6,9, but one problem with many of the biological markers used to assess muscle injury, including those used in animal studies, is that they usually do not correlate with the loss of force. Muscle damage is often defined within the context of the assay used to examine it and no one finding can account for the changes in contractility after injury. Since full contractile function can persist...

(All equipment is the same for mice and rats except for the footplate)
<ul>
	<li>BUD Value Line Cabinet (Newark, 06M4718)</li>
	<li>Multifunction l/O USB-6221M (National Instruments, 779808-01)</li>
	<li>Stepper motor controller (Newark, 16M4189)</li>
	<li>Stepper Motor (Newark, 16M4198)</li>
	<li>Strain Gauge Amplifier (Honeywell, Sensotec, DV-05)</li>
	<li>Torque Sensor (Honeywell, QWLC-8M)</li>
	<li>Foot plate and stabilization device (custom made, patent pending)</li>
</ul>

an in vivo rodent model of contraction-induced injury and non-invasive monitoring of recovery

Muscle strains are one of the most common complaints treated by physicians. A muscle injury is typically diagnosed from the patient history and physical exam alone, however the clinical presentation can vary greatly depending on the extent of injury, the patient's pain tolerance, etc. In patients with muscle injury or muscle disease, assessment of muscle damage is typically limited to clinical signs, such as tenderness, strength, range of motion, and more recently, imaging studies. Biological markers, such as serum creatine kinase levels, are typically elevated with muscle injury, but their levels do not always correlate with the loss of force production. This is even true of histological findings from animals, which provide a "direct measure" of damage, but do not account for all the loss of function. Some have argued that the most comprehensive measure of the overall health of the muscle in contractile force. Because muscle injury is a random event that occurs under a variety of biomechanical conditions, it is difficult to study. Here, we describe an in vivo animal model to measure torque and to produce a reliable muscle injury. We also describe our model for measurement of force from an isolated muscle in situ. Furthermore, we describe our small animal MRI procedure.

Watch this Scientific Journal Video about An in vivo Rodent Model of Contraction-induced Injury and Non-invasive Monitoring of Recovery at JoVE.com

An in vivo Rodent Model of Contraction-induced Injury and Non-invasive Monitoring of Recovery

An in vivo animal model of injury is described. The method takes advantage of the subcutaneous position of the fibular nerve. Velocity, timing of muscle activation, and arc of motion are all pre-determined and synchronized using commercial software. Post injury changes are monitored in vivo using MR imaging/spectroscopy.

An in vivo Rodent Model of Contraction-induced Injury and Non-invasive Monitoring of Recovery

Muscle strains are one of the most  common complaints treated by physicians. A muscle injury is typically diagnosed from the patient history and  physical ...

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13.7K Views.  University of Maryland School of Medicine. An in vivo animal model of injury is described. The method takes advantage of the subcutaneous position of the fibular nerve. Velocity, timing of muscle activation, and arc of motion are all pre-determined and synchronized using commercial software. Post injury changes are monitored in vivo using MR imaging/spectroscopy.

Video: An in vivo Rodent Model of Contraction-induced Injury and Non-invasive Monitoring of Recovery

Non-invasive Optical Measurement of Cerebral Metabolism and Hemodynamics in Infants

Perinatal brain injury remains a significant cause of infant mortality and morbidity, but there is not yet an effective bedside tool that can accurately screen for brain injury, monitor injury evolution, or assess response to therapy. The energy used by neurons is derived largely from tissue oxidative metabolism, and neural hyperactivity and cell death are reflected by corresponding changes in cerebral oxygen metabolism (CMRO2). Thus, measures of CMRO2 are reflective of neuronal viability and provide critical diagnostic information, making CMRO2 an ideal target for bedside measurement of brain health.
Brain-imaging techniques such as positron emission tomography (PET) and single-photon emission computed tomography (SPECT) yield measures of cerebral glucose and oxygen metabolism, but these techniques require the administration of radionucleotides, so they are used in only the most acute cases.
Continuous-wave near-infrared spectroscopy (CWNIRS) provides non-invasive and non-ionizing radiation measures of hemoglobin oxygen saturation (SO2) as a surrogate for cerebral oxygen consumption. However, SO2 is less than ideal as a surrogate for cerebral oxygen metabolism as it is influenced by both oxygen delivery and consumption. Furthermore, measurements of SO2 are not sensitive enough to detect brain injury hours after the insult 1,2, because oxygen consumption and delivery reach equilibrium after acute transients 3. We investigated the possibility of using more sophisticated NIRS optical methods to quantify cerebral oxygen metabolism at the bedside in healthy and brain-injured newborns. More specifically, we combined the frequency-domain NIRS (FDNIRS) measure of SO2 with the diffuse correlation spectroscopy (DCS) measure of blood flow index (CBFi) to yield an index of CMRO2 (CMRO2i) 4,5.
With the combined FDNIRS/DCS system we are able to quantify cerebral metabolism and hemodynamics. This represents an improvement over CWNIRS for detecting brain health, brain development, and response to therapy in neonates. Moreover, this method adheres to all neonatal intensive care unit (NICU) policies on infection control and institutional policies on laser safety. Future work will seek to integrate the two instruments to reduce acquisition time at the bedside and to implement real-time feedback on data quality to reduce the rate of data rejection.

We combined frequency-domain near-infrared spectroscopy measures of cerebral hemoglobin oxygenation with diffuse correlation spectroscopy measures of cerebral blood flow index to estimate an index of oxygen metabolism. We tested the utility of this measure as a bedside screening tool to evaluate the health and development of the newborn brain.

Perinatal brain injury remains a significant cause of infant mortality  and morbidity, but there is not yet an effective bedside tool that can  accurately ...

Non-invasive Assessment of Microvascular and Endothelial Function

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of cardiovascular disease. Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. Percent capillary recruitment is assessed by dividing the increase in capillary density induced by postocclusive reactive hyperemia (postocclusive reactive hyperemia capillary density minus baseline capillary density), by the maximal capillary density (observed during passive venous occlusion). Percent perfused capillaries represents the proportion of all capillaries present that are perfused (functionally active), and is calculated by dividing postocclusive reactive hyperemia capillary density by the maximal capillary density. Both percent capillary recruitment and percent perfused capillaries reflect the number of functional capillaries. The forearm blood flow (FBF) technique provides accepted non-invasive measures of endothelial function: The ratio FBFmax/FBFbase is computed as an estimate of vasodilation, by dividing the mean of the four FBFmax values by the mean of the four FBFbase values. Forearm vascular resistance at maximal vasodilation (FVRmax) is calculated as the mean arterial pressure (MAP) divided by FBFmax. Both the capillaroscopy and forearm techniques are readily acceptable to patients and can be learned quickly.
The microvascular and endothelial function measures obtained using the methodologies described in this paper may have future utility in clinical patient cardiovascular risk-reduction strategies. As we have published reports demonstrating that microvascular and endothelial dysfunction are found in initial stages of hypertension including prehypertension, microvascular and endothelial function measures may eventually aid in early identification, risk-stratification and prevention of end-stage vascular pathology, with its potentially fatal consequences.

Capillaroscopy is a non-invasive, relatively inexpensive methodology for directly visualizing the microcirculation. The forearm blood flow technique provides accepted non-invasive measures of endothelial function.

The authors have utilized capillaroscopy and forearm blood flow techniques to investigate the role of microvascular dysfunction in pathogenesis of ...

Transplantation into the Anterior Chamber of the Eye for Longitudinal, Non-invasive In vivo Imaging with Single-cell Resolution in Real-time

Intravital imaging has emerged as an indispensable tool in biological research. In the process, many imaging techniques have been developed to study different biological processes in animals non-invasively. However, a major technical limitation in existing intravital imaging modalities is the inability to combine non-invasive, longitudinal imaging with single-cell resolution capabilities. We show here how transplantation into the anterior chamber of the eye circumvents such significant limitation offering a versatile experimental platform that enables non-invasive, longitudinal imaging with cellular resolution in vivo. We demonstrate the transplantation procedure in the mouse and provide representative results using a model with clinical relevance, namely pancreatic islet transplantation. In addition to enabling direct visualization in a variety of tissues transplanted into the anterior chamber of the eye, this approach provides a platform to screen drugs by performing long-term follow up and monitoring in target tissues. Because of its versatility, tissue/cell transplantation into the anterior chamber of the eye not only benefits transplantation therapies, it extends to other in vivo applications to study physiological and pathophysiological processes such as signal transduction and cancer or autoimmune disease development.

Transplantation into the Anterior Chamber of the Eye for Longitudinal, Non-invasive In vivo Imaging with Single-cell Resolution in Real-time

A new approach combining intraocular transplantation and confocal microscopy enables longitudinal, non-invasive real-time imaging with single-cell resolution within grafted tissues in vivo. We demonstrate how to transplant pancreatic islets into the anterior chamber of the mouse eye.

Intravital imaging has emerged as an indispensable tool in biological research.  In the process, many imaging techniques have been developed to study ...

An Alkali-burn Injury Model of Corneal Neovascularization in the Mouse

Under normal conditions, the cornea is avascular, and this transparency is essential for maintaining good visual acuity. Neovascularization (NV) of the cornea, which can be caused by trauma, keratoplasty or infectious disease, breaks down the so called &lsquo;angiogenic privilege' of the cornea and forms the basis of multiple visual pathologies that may even lead to blindness. Although there are several treatment options available, the fundamental medical need presented by corneal neovascular pathologies remains unmet. In order to develop safe, effective, and targeted therapies, a reliable model of corneal NV and pharmacological intervention is required. Here, we describe an alkali-burn injury corneal neovascularization model in the mouse. This protocol provides a method for the application of a controlled alkali-burn injury to the cornea, administration of a pharmacological compound of interest, and visualization of the result. This method could prove instrumental for studying the mechanisms and opportunities for intervention in corneal NV and other neovascular disorders.

Neovascularization (NV) of the cornea can complicate multiple visual pathologies. Utilizing a controlled, alkali-burn injury model, a quantifiable level of corneal NV can be produced for mechanistic study of corneal NV and evaluation of potential therapies for neovascular disorders.

Under normal conditions, the cornea is avascular, and this transparency is essential for maintaining good visual acuity. Neovascularization (NV) of the ...

Murine Model for Non-invasive Imaging to Detect and Monitor Ovarian Cancer Recurrence

Epithelial ovarian cancer is the most lethal gynecologic malignancy in the United States. Although patients initially respond to the current standard of care consisting of surgical debulking and combination chemotherapy consisting of platinum and taxane compounds, almost 90% of patients recur within a few years. In these patients the development of chemoresistant disease limits the efficacy of currently available chemotherapy agents and therefore contributes to the high mortality. To discover novel therapy options that can target recurrent disease, appropriate animal models that closely mimic the clinical profile of patients with recurrent ovarian cancer are required. The challenge in monitoring intra-peritoneal (i.p.) disease limits the use of i.p. models and thus most xenografts are established subcutaneously. We have developed a sensitive optical imaging platform that allows the detection and anatomical location of i.p. tumor mass. The platform includes the use of optical reporters that extend from the visible light range to near infrared, which in combination with 2-dimensional X-ray co-registration can provide anatomical location of molecular signals. Detection is significantly improved by the use of a rotation system that drives the animal to multiple angular positions for 360 degree imaging, allowing the identification of tumors that are not visible in single orientation. This platform provides a unique model to non-invasively monitor tumor growth and evaluate the efficacy of new therapies for the prevention or treatment of recurrent ovarian cancer.

We describe a non-invasive animal imaging platform that allows the detection, quantification, and monitoring of ovarian cancer growth and recurrence. This intra-peritoneal xenograft model mimics the clinical profile of patients with ovarian cancer.

Epithelial ovarian cancer is the most lethal gynecologic malignancy in the United States. Although patients initially respond to the current standard of ...

Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

Several in vivo pre-clinical studies in Polycystic Kidney Disease (PKD) utilize orthologous rodent models to identify and study the genetic and molecular mechanisms responsible for the disease, and are very convenient for rapid drug screening and testing of promising therapies. A limiting factor in these studies is often the lack of efficient non-invasive methods for sequentially analyzing the anatomical and functional changes in the kidney. Magnetic resonance imaging (MRI) is the current gold standard imaging technique to follow autosomal dominant polycystic kidney disease (ADPKD) patients, providing excellent soft tissue contrast and anatomic detail and allowing Total Kidney Volume (TKV) measurements.A major advantage of MRI in rodent models of PKD is the possibility for in vivo imaging allowing for longitudinal studies that use the same animal and therefore reducing the total number of animals required. In this manuscript, we will focus on using Ultra-high field (UHF) MRI to non-invasively acquire in vivo images of rodent models for PKD. The main goal of this work is to introduce the use of MRI as a tool for in vivo phenotypical characterization and drug monitoring in rodent models for PKD.

Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

The use of ultra-high field MRI as a non-invasive way to obtain phenotypic information of rodent models for polycystic kidney disease and to monitor interventions is described. Compared with the traditional histological approach, MRI images can be acquired in vivo, allowing for longitudinal follow-up.

Several in vivo pre-clinical studies in Polycystic Kidney Disease (PKD) utilize orthologous rodent models to identify and study the genetic and molecular ...

Non-invasive Assessments of Subjective and Objective Recovery Characteristics Following an Exhaustive Jump Protocol

Fast recovery after strenuous exercise is important in sports and is often studied via cryotherapy applications. Cryotherapy has a significant vasoconstrictive effect, which seems to be the leading factor in its effectiveness. The resulting enhanced recovery can be measured by using both objective and subjective parameters. Two commonly measured subjective characteristics of recovery are delayed-onset muscle soreness (DOMS) and ratings of perceived exertion (RPE). Two important objective recovery characteristics are countermovement jump (CMJ) performance and peak power output (PPO). Here, we provide a detailed protocol to induce muscular exhaustion of the frontal thighs with a self-paced, 3 x 30 countermovement jump protocol (30-s rest between each set). This randomized controlled trial protocol explains how to perform local cryotherapy cuff application (+ 8 &#176;C for 20 min) and thermoneutral cuff application (+ 32 &#176;C for 20 min) on both thighs as two possible post-exercise recovery modalities. Finally, we provide a non-invasive protocol to measure the effects of these two recovery modalities on subjective (i.e., DOMS of both frontal thighs and RPE) and objective recovery (i.e., CMJ and PPO) characteristics 24, 48, and 72 h post-application. The advantage of this method is that it provides a tool for researchers or coaches to induce muscular exhaustion, without using any expensive devices; to implement local cooling strategies; and to measure both subjective and objective recovery, without using invasive methods. Limitations of this protocol are that the 30 s rest period between sets is very short, and the cardiovascular demand is very high. Future studies may find the assessment of maximum voluntary contractions to be a more sensitive assessment of muscular exhaustion compared to CMJs.

This protocol describes the procedure for non-invasive recovery assessments during a 72 h recovery period. This protocol induces muscular exhaustion of the frontal thighs using countermovement jumps and uses either local cold-cuff or thermoneutral-cuff application as a recovery modality.

Fast recovery after strenuous exercise is important in sports and is often studied via cryotherapy applications. Cryotherapy has a significant ...

Non-invasive Assessment of Dorsiflexor Muscle Function in Mice

Assessment of skeletal muscle contractile function is an important measurement for both clinical and research purposes. Numerous conditions can negatively affect skeletal muscle. This can result in a loss of muscle mass (atrophy) and/or loss of muscle quality (reduced force per unit of muscle mass), both of which are prevalent in chronic disease, muscle-specific disease, immobilization, and aging (sarcopenia). Skeletal muscle function in animals can be evaluated by a range of different tests. All tests have limitations related to the physiological testing environment, and the selection of a specific test often depends on the nature of the experiments. Here, we describe an in vivo, non-invasive technique involving a helpful and easy assessment of force frequency-curve (FFC) in mice that can be performed on the same animal over time. This permits monitoring of disease progression and/or efficacy&#160;of a potential therapeutic treatment.

Measurement of rodent skeletal muscle contractile function is a useful tool that can be used to track disease progression as well as efficacy of therapeutic intervention. We describe here the non-invasive, in vivo assessment of the dorsiflexor muscles that can be repeated over time in the same mouse.

Assessment of skeletal muscle contractile function is an important measurement for both clinical and research purposes. Numerous conditions can negatively ...

An Intact Pericardium Ischemic Rodent Model

This protocol has shown that the pericardium and its contents play an essential anti-fibrotic role in the ischemic rodent model (coronary ligation to induce myocardial injury). The majority of pre-clinical myocardial infarction models require the disruption of pericardial integrity with loss of the homeostatic cellular milieu. However, recently a methodology has been developed by us to induce myocardial infarction, which minimizes pericardial damage and retains the heart&#39;s resident immune cell population. An improved cardiac functional recovery in mice with an intact pericardial space following coronary ligation has been observed. This method provides an opportunity to study inflammatory responses in the pericardial space following myocardial infarction. Further development of the labeling techniques can be combined with this model to understand the fate and function of pericardial immune cells in regulating the inflammatory mechanisms that drive remodeling in the heart, including fibrosis.

This protocol outlines the steps for inducing myocardial infarction in mice while preserving the pericardium and its contents.

This protocol has shown that the pericardium and its contents play an essential anti-fibrotic role in the ischemic rodent model (coronary ligation to ...

Improved Rodent Model of Myocardial Ischemia and Reperfusion Injury

Myocardial ischemia and reperfusion injury (MIRI), induced by coronary heart disease (CHD), causes damage to the cardiomyocytes. Furthermore, evidence suggests that thrombolytic therapy or primary percutaneous coronary intervention (PPCI) does not prevent reperfusion injury. There is still no ideal animal model for MIRI. This study aims to improve the MIRI model in rats to make surgery easier and more feasible. A unique method for establishing MIRI is developed by using a soft tube during a key step of the ischemic period. To explore this method, thirty rats were randomly divided into three groups: sham group (n = 10); experimental model group (n = 10); and existing model group (n = 10). Findings of triphenyltetrazolium chloride staining, electrocardiography, and percent survival are compared to determine the accuracies and survival rates of the operations. Based on the study results, it has been concluded that the improved surgery method is associated with a higher survival rate, elevated ST-T segment, and larger infarct size, which is expected to mimic the pathology of MIRI better.

Myocardial ischemia-reperfusion model of rat heart is improved by using a self-made retractor, polyvinyl chloride tube, and a unique knotting method. Electrocardiogram, triphenyltetrazolium chloride and histological staining, and percent survival analysis results showed that the improved model group has higher success and survival rates than the already existing model group.

Myocardial ischemia and reperfusion injury (MIRI), induced by coronary heart disease (CHD), causes damage to the cardiomyocytes. Furthermore, evidence ...