Bioclimatic thermal stress indices and their relationships with andrological characteristics in hair rams.

This study evaluated relationships among reproductive parameters and the bioclimatic indices: temperature and humidity index (THI), equivalent temperature index (ETI), black globe temperature and humidity index (BGTHI), and thermal comfort index (TCI), during the first 45 days of spermatogenesis (SP-45) and during the 15 days of sperm transit through the epididymis (STP-15) that preceded the reproductive assessments (ReA). Such information is useful in determining the optimal breeding season in Northeast Brazil. Santa Inês rams (n = 25) underwent two ReA in three periods of the year (D-P = dry; R-P = rainy and RD-P = rainy/dry transition), and the bioclimatic indices were calculated at the corresponding SP-45 and STP-15 timepoints prior to each ReA. Sperm kinetic parameters in D-P were depressed compared to R-P and RD-P (P < 0.05). The index values had an antagonistic relationship with most parameters and regression analysis demonstrated that the BGTHI and the TCI had a negative association with the progressive motility, curvilinear, straight line, and average path velocities, and a positive association with slow sperm in the ejaculate in SP-45 and STP-15 phases (P < 0.01). Semen quality kinetics is affected throughout the year by the environment and it is apparent that it is impaired in D-P and better in R-P and RD-P seasons. The BGTHI and TCI measured in the sperm production phase classified the environment most coherently and presented better association with the behavior of sperm kinetics.

Cryopreservation of turkey semen: effect of breeding line and freezing method on post-thaw sperm quality, fertilization, and hatching.

Cryopreservation methods for poultry semen are not reliable for germplasm preservation, especially for turkeys, where fertility rates from frozen/thawed semen are particularly low. The objective was to evaluate cryopreservation methods for effectiveness in promoting cryosurvival and post-thaw function of sperm from five turkey lines: one commercial line and four research (RBC1; E; RBC2; F) lines from Ohio State University (OSU). The model for cryopreservation was set up as a 2×2×2×5 design for cryoprotectant (glycerol or dimethylacetamide (DMA)), cryopreservation medium (Lake or ASG), method of dilution (fixed dilution volume versus fixed sperm concentration) and turkey line, respectively. The final cryoprotectant concentrations were 11% glycerol or 6% DMA. Thawed sperm were evaluated for plasma membrane integrity and quality, motility, acrosome integrity and, after artificial insemination, for egg fertility and hatchability. Commercial turkey hens were used for all fertility trials, regardless of semen source. Turkey sperm frozen with glycerol exhibited higher membrane integrity and membrane quality upon thawing than turkey sperm frozen with DMA although no differences in total motility, and only minimal differences in progressive motility, were detected among the eight cryopreservation treatments. Within line, fertility was affected by cryoprotectant, medium and dilution method, where the overall highest percentages of fertile, viable embryos (Day 7) occurred for the DMA/ASG/fixed sperm concentration method, while high percentages (15.8-31.5%) of fertile, non-viable embryos (Day 1-6) were observed for multiple cryopreservation methods, including two glycerol treatments. From a single insemination, the duration of true and viable fertility in all lines was 10-13 weeks and 9-10 weeks, respectively. The duration of hatchability was 4-6 weeks after insemination for four of the turkey lines. The highest percentage of viable embryos was observed for the commercial line (9.5±2.4%), followed by the E line (5.3±1.3%), F line (3.7±2.0%) and RBC2 line (2.6±0.8%). For the RBC1 line, there was 100% embryonic death by Day 6 of incubation. Overall, better fertility results were obtained with the cryoprotectant DMA, the ASG diluent and fixed sperm concentration. However, the applicability of this method for preserving semen from research populations may be line dependent.

Direct in vitro propagation of avian germ cells from an embryonic gonad biorepository.

Direct introduction of cryopreserved embryonic gonadal germ cells (GGC) into a sterile chicken surrogate host to reconstitute a chicken breed has been demonstrated as a feasible approach for preserving and utilizing chicken genetic resources. This method is highly efficient using male gonads; however, a large number of frozen female embryonic gonads is needed to provide sufficient purified GGC for the generation of fertile surrogate female hosts. Applying this method to indigenous chicken breeds and other bird species is difficult due to small flock numbers and poor egg production in each egg laying cycle. Propagating germ cells from the frozen gonadal tissues may be a solution for the biobanking of these birds. Here, we describe a simplified method for culture of GGC from frozen embryonic 9.5 d gonads. At this developmental stage, the germ cells are autonomously shed into medium, yielding hundreds to thousands of mitosis-competent germ cells. The resulting cultures of GGC have over 90% purity, uniformly express SSEA-1 and DAZL antigens and can re-colonize recipient's gonads. The GGC recovery rate from frozen gonads are 42% to 100%, depending on length of cryopreservation and the breed or line of chickens. Entire chicken embryos can also be directly cryopreserved for later gonadal isolation and culture. This storage method is a supplementary approach to safeguard local indigenous chicken breeds bearing valuable genetic traits and should be applicable to the biobanking of many bird species.

Incorporation of Biotechnologies into Gene Banking Strategies to Facilitate Rapid Reconstitution of Populations.

National animal gene banks that are responsible for conserving livestock, poultry, and aquatic genetic resources need to be capable of utilizing a broad array of cryotechnologies coupled with assisted reproductive technologies to reconstitute either specific animals or populations/breeds as needed. This capability is predicated upon having sufficient genetic diversity (usually encapsulated by number of animals in the collection), units of germplasm or tissues, and the ability to reconstitute animals. While the Food and Agriculture Organization of the United Nations (FAO 2012, 2023) developed a set of guidelines for gene banks on these matters, those guidelines do not consider applications and utilization of newer technologies (e.g., primordial germ cells, cloning from somatic cells, embryo transfer, IVF, sex-sorted semen), which can radically change how gene banks collect, store, and utilize genetic resources. This paper reviews the current status of using newer technologies, explores how gene banks might make such technologies part of their routine operations, and illustrates how combining newer assisted reproductive technologies with older approaches enables populations to be reconstituted more efficiently.

Evaluation of boar and bull sperm capacitation and the acrosome reaction using flow cytometry.

Flow cytometry can be used to evaluate many sperm attributes and Dr. Duane Garner was influential in developing assays to understand sperm physiology and function. We review some of Dr. Garner's work and describe experiments that evaluate sperm capacitation using Dr. Garner's philosophy. In exploratory experiments, boar sperm were cryopreserved in lactose egg yolk (LEY) or Beltsville Freezing Extender 5 (BF5) and incubated in one capacitating medium. In another experiment, frozen-thawed bull sperm were incubated in TALP-Ca or CFDM1 capacitating media. In both experiments, sperm viability and capacitation were evaluated using multiple probes. Boar sperm frozen in LEY had greater survival rates (38%) than sperm frozen in BF5 (22%; P < 0.05) but did not capacitate as effectively as sperm in BF5 (P < 0.05). In Experiment 2, bull sperm survived to a greater extent when incubated in TALP-Ca than in CFDM1 (P < 0.05) and had greater capacitation for most parameters (P < 0.05). Of particular interest, 77% of sperm incubated in TALP-Ca had activated second messenger systems involved in capacitation, compared with < 5% of sperm incubated in CFDM1. The results indicate different freezing and capacitating media induce different responses to sperm capacitation and functions. If only sperm viability and acrosomal integrity were evaluated, these results would be interpreted very differently. Dr. Garner's philosophy of evaluating multiple sperm parameters was an impetus to determine unique treatment differences which help in understanding sperm capacitation, and design further experiments to determine how media content causes sperm physiology differences.

Target lesion revascularization after wingspan: assessment of safety and durability.

BACKGROUND AND PURPOSE: In-stent restenosis (ISR) occurs in approximately one-third of patients after the percutaneous transluminal angioplasty and stenting of intracranial atherosclerotic lesions with the Wingspan system. We review our experience with target lesion revascularization (TLR) for ISR after Wingspan treatment. METHODS: Clinical and angiographic follow-up results were recorded for all patients from 5 participating institutions in our US Wingspan Registry. ISR was defined as >50% stenosis within or immediately adjacent (within 5 mm) to the implanted stent and >20% absolute luminal loss. RESULTS: To date, 36 patients in the registry have experienced ISR after percutaneous transluminal angioplasty and stenting with Wingspan. Of these patients, 29 (80.6%) have undergone TLR with either angioplasty alone (n=26) or angioplasty with restenting (n=3). Restenting was performed for in-stent dissections that occurred after the initial angioplasty. Of the 29 patients undergoing TLR, 9 required >/=1 interventions for recurrent ISR, for a total of 42 interventions. One major complication, a postprocedural reperfusion hemorrhage, was encountered in the periprocedural period (2.4% per procedure; 3.5% per patient). Angiographic follow-up is available for 22 of 29 patients after TLR. Eleven of 22 (50%) demonstrated recurrent ISR at follow-up angiography. Nine patients have undergone multiple retreatments (2 retreatments, n=6; 3 retreatments, n=2; 4 retreatments, n=1) for recurrent ISR. Nine of 11 recurrent ISR lesions were located within the anterior circulation. The mean age for patients with recurrent anterior circulation ISR was 57.9 years (vs 81 years for posterior circulation ISR). CONCLUSIONS: TLR can be performed for the treatment of intracranial Wingspan ISR with a relatively high degree of safety. However, the TLR results are not durable in approximately 50% of patients, and multiple revascularization procedures may be required in this subgroup.

Stent/coil treatment of very large and giant unruptured ophthalmic and cavernous aneurysms.

BACKGROUND: Treatment of VLGUIA remains a challenge. To reduce mass effect and achieve complete occlusion, open surgery has been our favored treatment. However, endovascular therapy is preferred for lesions in the cavernous sinus or for older patients with complicating medical problems. The goal of this study is to investigate outcome of stent and/or coil treatment of VLGUIA. METHODS: Beginning in 2002, the neuroform stent has been available to the University of Texas Southwestern Medical Center in Dallas. Since then until 2006, 15 patients were treated for VLGUIA with stenting and/or coiling at this institution. These 15 patients were used for a retrospective analysis in this study. RESULTS: Median patient age was 65 years, median aneurysm size was 27 mm (20-37 mm), and median follow-up time was 22 months. Eight aneurysms were localized in the cavernous sinus and 7 at the ophthalmic segment of the internal carotid artery. Four aneurysms were completely occluded (100%); 3 aneurysms, nearly complete (90%-99%); and 8 aneurysms, partial (<90% occlusion). Twelve patients required retreatment. Final GOS was 1 (good recovery) in 11 patients, 2 (moderate disability) in 3 patients, and 3 (severely disabled) in 1 patient. No patient died or deteriorated. CONCLUSIONS: Stent/coil management of VLGUIA is constantly evolving. Current treatment results are promising, with very low morbidity/mortality. Disadvantage is the frequent persistence of residual aneurysm.

Swine gene banking: a quality control perspective on collection, and analysis of samples for a national repository.

The National Animal Germplasm Program (NAGP) is developing a national repository for germplasm (semen, oocytes, embryos, blood, DNA, tissue) for all agricultural species in the US. Currently, the swine collection consists of 127,479 samples from 886 boars representing 20 major, minor and composite populations. Cryopreservation per se is not an impediment to program success. Rather, the greatest difficulties encountered are in determining the quality of the samples pre- and post-thaw. Robust, broadly applicable, and cost effective quality control methodologies need to be developed and implemented. This overview of the NAGP will discuss the approaches used for cryopreserving boar semen samples, overcoming the challenges of assessing sample quality, and moving toward a quality control strategy.

The effects of gelatin supplementation prior to cooling on ram semen quality and fertility.

The physical and chemical characteristics of gelatin have been used to justify its inclusion in extenders to preserve the sperm quality and improve results of cervical artificial insemination with cooled semen. The objective of this study was to evaluate the effects of gelatin supplementation in cooling extender on the quality and fertility of ram semen stored at 5°C. Semen samples (n = 24) of Santa Inês rams (n = 6) were diluted in Glycine-Yolk-Milk extender without (control) or with 1.5% of gelatin. The samples were loaded into 0.25 mL straws, cooled to 5°C and stored vertically for 48 and 72 hours. Sample quality was evaluated using straw homogeneity tests based on pH, osmolality and the proportion of spermatozoa (PS) in both upper and lower segments of straws (US and LS), analyses of sperm motility, plasma and acrosomal membrane integrity, and by fertility after artificial insemination. Differences between the US and LS of straws were found for pH and PS (%). They were significant only in the control group at both times: pH - 5.96 vs. 5.71 at 48 h and 6.13 vs. 5.89 at 72 h; PS - 21.66 vs. 78.34 at 48 h and 20.87 vs. 79.13 at 72 h. Storage in gelatin had very little, to no effect on the sperm kinetics or on the sperm membrane integrity evaluations. The addition of gelatin to the extender did not affect the pregnancy rate which ranged from 4.4 to 26.1%. We conclude that gelatin is effective in maintaining the physical and chemical homogeneity of the semen samples. Further research is needed in order to optimize the use of gelatin supplementation and elucidate any potential benefits.

Use of a pressure sensing sheath: comparison with standard means of blood pressure monitoring in catheterization procedures.

PURPOSE: Monitoring of blood pressure (BP) during procedures is variable, depending on multiple factors. Common methods include sphygmomanometer (BP cuff), separate radial artery catheterization, and side port monitoring of an indwelling sheath. Each means of monitoring has disadvantages, including time consumption, added risk, and signal dampening due to multiple factors. We sought an alternative approach to monitoring during procedures in the catheterization laboratory. METHODS: A new technology involving a 330 µm fiberoptic sensor embedded in the wall of a sheath structure was tested against both radial artery catheter and sphygmomanometer readings obtained simultaneous with readings recorded from the pressure sensing system (PSS). Correlations and Bland-Altman analysis were used to determine whether use of the PSS could substitute for these standard techniques. RESULTS: The results indicated highly significant correlations in systolic, diastolic, and mean arterial pressures (MAP) when compared against radial artery catheterization (p<0.0001), and MAP means differed by <4%. Bland-Altman analysis of the data suggested that the sheath measurements can replace a separate radial artery catheter. While less striking, significant correlations were seen when PSS readings were compared against BP cuff readings. CONCLUSIONS: The PSS has competitive functionality to that seen with a dedicated radial artery catheter for BP monitoring and is available immediately on sheath insertion without the added risk of radial catheterization. The sensor is structurally separated from the primary sheath lumen and readings are unaffected by device introduction through the primary lumen. Time delays and potential complications from radial artery catheterization are avoided.

Visualization of sacral nerve roots via percutaneous intraspinal navigation (PIN).

A percutaneous technique for visualizing sacral nerve roots is described. A fiberscope was inserted into the subarachnoid space through a sheath that was inserted via a percutaneous lumbar puncture. The sacral nerve roots were identified with endoscopic visualization and x-ray fluoroscopy localization of the endoscope. These images were compared with those obtained from a videoscope, which revealed better imaging. Specific sacral nerve roots can be identified by using a combination of endoscopy and x-ray fluoroscopy. This technique may enable minimally invasive interventions such as lysis of adhesions, arachnoid cyst decompression, selective dorsal rhizotomy, and more selective and precise nerve stimulation electrode placement.

Percutaneous intraspinal navigation for access to the subarachnoid space: use of another natural conduit for neurosurgical procedures.

OBJECT: The purpose of this paper was to demonstrate the usefulness of various fiberoptic endoscopes for percutaneous intraspinal navigation of the spinal canal, posterior fossa, and ventricular system. METHODS: Fresh, unembalmed cadavers were used, in which lumbar punctures were made for access to the subarachnoid space (in the case of larger [3.8- and 5-mm-diameter] endoscopes, small laminotomies were performed). Static and video images of pertinent structures were acquired for comparison among devices. Endoscopes were compared for their maneuverability, durability, field of view, and image quality. Seven sizes and types of endoscopes were considered. Overall, the devices offering a tip-deflecting mechanism were superior in maneuverability. Endoscopes in which a charged couple display chip was used at the tip of the scope for image acquisition offered improved image quality and field of view. Larger scopes, although more durable, were more rigid and may be limited in application. Multiple images from multiple devices are presented. CONCLUSIONS: Percutaneous intraspinal navigation offers a promising neurosurgical approach to the spinal canal, the posterior fossa, and the ventricular system. Concerns regarding safety, management of complications, and the lack of adjunctive tools for intervention through the endoscopes or for use under fluoroscopic guidance represent areas that warrant further investigation and development.

Membrane modification strategies for cryopreservation.

Cell membranes can be modified using cyclodextrins loaded with lipids or unilamellar liposomes. Lipid choice can greatly influence the organization of the targeted membrane and result in a cell that is more capable of surviving cryopreservation due to altered membrane-phase transition properties or membrane reorganization that may alter the normal physiologic processes of the treated cell. The protocols described here explain the preparation of the cyclodextrins and liposomes, impact of the amount and type of lipids, and general principles for treating cells using either of these technologies.

Interventional MR imaging with an endospinal imaging coil: preliminary results with anatomic imaging of the canine and cadaver spinal cord.

Percutaneous intraspinal navigation (PIN) is a new minimally invasive approach to the subarachnoid space. Using conventional radiographic fluoroscopy, entrance is gained to the lumbar subarachnoid space, allowing navigation throughout the spinal canal. Using an antenna/guidewire introduced via PIN, we performed endospinal MR imaging of the thoracic spinal cord in a cadaver and canine subject. Comparison images were obtained with an optimal surface coil. PIN allows endospinal MR imaging of the spinal cord, providing significant signal-to-noise ratio gains over conventional imaging.

Percutaneous intraspinal navigation: feasibility study of a new and minimally invasive approach to the spinal cord and brain in cadavers.

We describe a percutaneous approach for cerebral surgical access. After lumbar puncture, the spinal subarachnoid space was traversed by using standard angiographic guidewire techniques until the introducer catheters were in the intracranial space. Under fluoroscopic guidance, the intracranial subarachnoid space was navigated, and the ventricular system entered. Subarachnoid placement was confirmed with contrast-enhanced digital angiography. Placement anterior to the brain stem was confirmed in both cadavers during dissection, and spinal navigation without cord damage from the anterior or posterior approach was confirmed in one. Percutaneous intraspinal navigation is a new route of access for cerebrospinal surgery that has many potential applications.

Stent-coil treatment of a distal internal carotid artery dissecting pseudoaneurysm on a redundant loop by use of a flexible, dedicated nitinol intracranial stent.

Treatment of dissecting pseudoaneurysms of the distal cervical internal carotid artery with preservation of the parent artery by using stents or coils has become routine. Tortuosity remains a significant obstacle to successful endovascular treatment in some cases. We report the use of a stent-coil technique to treat a nonhealing dissecting pseudoaneurysm and associated stenosis with anatomic preservation of a redundant loop involving the stented arterial segment. This was accomplished by using a Neuroform dedicated intracranial stent.

Percutaneous translumbar spinal cord compression injury in a dog model that uses angioplasty balloons: MR imaging and histopathologic findings.

BACKGROUND AND PURPOSE: Previous animal models for spinal cord injury required laminectomy and exposure of the spinal cord to create direct trauma, compromising imaging by both surgical artifact and the nature of the production of the injury. Our purpose was to study a model that uses percutaneous intraspinal navigation with an angioplasty balloon, providing a controlled degree of spinal cord compression and allowing improved MR imaging of spinal cord injury. METHODS: Nine mongrel dogs were studied. MR images were obtained of six dogs after technique development in three dogs. Angioplasty balloons measuring 7 or 4 mm in diameter and 2 cm in length were placed in the midthoracic subarachnoid space. Imaging was performed by using a 1.5-T MR imaging unit before and after balloon inflation. The balloon was inflated within 5 seconds and deflated after 30 minutes. T1- and T2-weighted and contrast-enhanced images were acquired. Spinal cords were submitted for pathologic examination. RESULTS: All four animals with 7-mm balloons experienced hemorrhage, and three had axonal injury revealed by histopathologic examination. One of two animals with 4-mm balloons experienced no injury, and one had axonal injury without hemorrhage. Regional parenchymal enhancement was seen in two of the animals with 7-mm balloons. CONCLUSION: This percutaneous spinal cord injury model results in a graduating degree of injury. It differs from previous techniques by avoiding surgical exposure and the associated artifacts, yet it offers histopathologic findings similar to those of human spinal cord injury. The canine spinal cord is amenable to MR imaging with clinical imaging units. Further evaluations with various durations of compression and various balloon sizes are warranted.

MR-guided catheter navigation of the intracranial subarachnoid space.

Percutaneous intraspinal navigation (PIN) is a new minimally invasive approach to the CNS. The authors studied the utility of MR-guided intracranial navigation following access to the subarachnoid compartment via PIN. The passive tracking technique was employed to visualize devices during intracranial navigation. Under steady-state free precession (SSFP) MR-guidance a microcatheter-microguidewire was successfully navigated to multiple brain foci in two cadavers. SSFP MR fluoroscopy possesses adequate contrast and temporal resolution to allow MR-guided intracranial navigation.

Percutaneous translumbar spinal cord compression injury in dogs from an angioplasty balloon: MR and histopathologic changes with balloon sizes and compression times.

BACKGROUND AND PURPOSE: Our previous model of spinal cord injury (SCI) included six dogs undergoing 30-minute compression with a balloon in the subarachnoid space. We determined whether various balloon sizes and compression times creates a gradation of injuries. METHODS: In 17 dogs (including our original six), angioplasty balloons 2, 4, or 7 mm in diameter (2 cm long) were inflated at T6 for 30, 120, or 240 minutes. T1- and T2-weighted, gadolinium-enhanced, and short-tau inversion recovery (STIR) MR images were obtained at 1.5 T. Spinal canal occlusion (SCO) was measured as balloon area-spinal cord area. Hematoxylin-eosin and beta amyloid precursor protein staining were performed to demonstrate hemorrhage and axonal injury, respectively. Injuries were scored as mild, moderate, or severe. Trends were assessed with one-way analysis of variance. RESULTS: SCO was 12.5-20% for 2-mm balloons, 28-56% for 4 mm, and 62-82% for 7 mm. No abnormalities were seen with SCO <30%. T1- and T2-weighted images had the poorest diagnostic performance; STIR images were best for predicting hemorrhage and axonal injury. Hemorrhage was demonstrated more frequently than was axonal injury. SCO (P < .0001) and hemorrhage (P = .002) significantly increased with balloon size. Longer inflation times tended to increase injuries for a given size, but differences were not significant. CONCLUSION: Compression injuries depended on the level of SCO. The compression times tested had less effect than the degree of compression. The value of 1.5-T MR imaging varied with the sequence and improved with contrast enhancement. STIR images showed SCIs not otherwise detected.