Immature Pigs Research Articles

The Efficacy of Terlipressin versus Adrenaline in Swine Cardiopulmonary Resuscitation

The objective of the present study was to evaluate the efficacy of terlipressin (TP) vs. adrenaline (ADR) in increasing coronary perfusion pressure (CPP) and return of spontaneous circulation (ROSC) in swine CPR. Under anesthesia with ketamine/thiopental, ventricular fibrillation was induced in 44 female immature pigs, remaining unassisted for 10 minutes, followed by 2 minutes of manual CPR (100 compression/10 ventilations/min with air). Animals were, then, divided into four groups: 1) ADR (45μg.kg(-1)); 2) saline-placebo (10mL); 3) TP 20μg.kg(-1)); and TP (20μg.kg(-1)) + ADR (45μg.kg(-1)). Defibrillation was performed after 2 minutes, observing surviving animals for a 30-minute period. Electrocardiogram, systemic BP, DBP, and PetCO(2) were monitored continuously. Terlipressin did not differ from placebo regarding the effects on CPP, with low rates of ROSC in both groups (1/11 vs. 2/11; p=NS). Adrenaline increased CPP from 13±12 to 54±15mmHg (p<0.0001), similar effect to TP + ADR (from 21±10 to 45±13mmHg; p<0.0001), with high rates of ROSC/survivors in both groups (10/11 vs. 9/11, respectively). Among survivors, greater MAP was observed in the TP + ADR group vs. ADR (105±19mmHg vs. 76±21mmHg; p=0.0157) groups. Adrenaline and TP + ADR were effective on maintaining CPP/ROSC in this experimental model, but isolated TP did not differ from placebo. However, in surviving animals in the TP + ADR group, greater hemodynamic stability was observed after ROSC, suggesting that TP can be a useful medication in the management of post-CPR hypotension.

Spinal growth modulation using a novel intravertebral epiphyseal device in an immature porcine model

Fusionless growth modulation is an attractive alternative to conventional treatments of idiopathic scoliosis. To date, fusionless devices achieve unilateral growth modulation by compressing the intervertebral disc. This study explores a device to control spinal alignment and vertebral morphology via growth modulation while excluding the disc in a porcine model. A device that locally encloses the vertebral growth plate exclusive of the disc was introduced anteriorly over T5-T8 in four immature pigs (experimental) while three underwent surgery without instrumentation (sham) and two were selected as controls. Bi-weekly coronal and lateral radiographs were taken over the 12-week follow-up to document vertebral morphology and spinal alignment modifications via an inverse approach (creation of deformity). All animals completed the experiment with no postoperative complications. Control and sham groups showed no significant changes in spinal alignment. Experimental group achieved a final coronal Cobb angle of 6.5°±3.5° (constrained to the four instrumented levels) and no alteration to the sagittal profile was observed. Solely the experimental group ended with consistent vertebral wedging of 4.1°±3.6° amounting to a cumulative wedging of up to 25° and a concurring difference in left/right vertebral height of 1.24±1.86mm in the coronal plane. The proposed intravertebral epiphyseal device, for the early treatment of progressive idiopathic scoliosis, demonstrated its feasibility by manipulating spinal alignment through the realization of local growth modulation exclusive of the intervertebral disc.

Effects of age and platelet‐rich plasma on ACL cell viability and collagen gene expression

Platelet-rich plasma (PRP) has shown in vivo potential to stimulate anterior cruciate ligament (ACL) healing at early time points in large animal models. However, in animal models, the healing potential of the ACL is dependent on animal age. In this study, we hypothesized that there are age-dependent differences in ACL cell metabolism, collagen gene expression, and the ability of the cells to respond to growth factors in PRP. To test this hypothesis, ACL cells were obtained from skeletally immature, adolescent and adult pigs, and cultured in a collagen type I hydrogel with or without PRP for 14 days. When cultured in collagen-only hydrogel, ACL cells from adult pigs had a 19% lower apoptotic rate as compared to immature pigs (p = 0.001) and a 25% higher cellular metabolic activity as compared to adolescent pigs (p = 0.006). The addition of PRP to the collagen hydrogel resulted in a significantly increased cellular metabolic activity, reduced apoptotic rate, and stimulation of collagen production in the cells from the immature and adolescent animals (p < 0.05 for all comparisons) but had less effect on adult cells. These findings suggest that skeletal maturity may influence ACL cells' metabolic activity, apoptosis, collagen production, and response to PRP.

Local Administration of Ibandronate and Bone Morphogenetic Protein-2 After Ischemic Osteonecrosis of the Immature Femoral Head

Bisphosphonate therapy has been shown to preserve the osteonecrotic femoral head in experimental and short-term clinical studies. However, a lack of new bone formation within the preserved femoral head due to the inhibition of bone remodeling is a concern. The purpose of this investigation was to determine if combined therapy consisting of ibandronate and bone morphogenetic protein-2 (BMP-2) can preserve the shape of the femoral head and stimulate new bone formation in an immature animal model of ischemic osteonecrosis. Ischemic osteonecrosis was surgically induced in immature pigs. Four groups were studied: normal, treated with saline solution, treated with ibandronate, and treated with both ibandronate and BMP-2 (the ibandronate + BMP-2 group). The animals were killed eight weeks after surgery. Radiographic, histological, and histomorphometric assessments were performed. Radiographic assessment showed better preservation of the femoral head shape-i.e., a 54% (CI [95% confidence interval]: 22%, 86%) higher mean epiphyseal quotient-in the ibandronate + BMP-2 group than in the saline group. Histological assessment showed increased trabecular bone in the ibandronate + BMP-2 group as compared with that in the saline group. The mean values for trabecular bone volume, thickness, and number and for osteoblast surface were an average of 400% (CI: 242%, 558%), 212% (CI: 166%, 259%), 71% (CI: 6%, 137%), and 2402% (CI: 2113%, 2693%) higher, respectively, in the ibandronate + BMP-2 group than in the saline group. The osteoclast number was significantly reduced in the ibandronate + BMP-2 group compared with that in the saline group (-59% [CI: -75%, -42%]). The mean osteoblast surface value in the ibandronate + BMP-2 group was significantly higher (2567% [CI: 2258%, 2877%]) than that in the ibandronate group. Heterotopic ossifications were present in the capsule of the hip joint in the ibandronate + BMP-2 group. A combination of ibandronate and BMP-2 decreased femoral head deformity while stimulating bone formation in an immature animal model of ischemic osteonecrosis.

Reactivity of intraosseous femoral head arteries after long term glucocorticoid treatment: An experimental study in immature pigs

In the pathogenesis of steroid induced femoral head necrosis only intra- and extravascular factors have been discussed. Vasoreactivity of intraosseous femoral head arteries has not previously been investigated. This study investigates the effect of long term glucocorticoid treatment on reactivity of intraosseous femoral head arteries in a porcine model. From 24 immature female Danish Landrace pigs from 12 litters, 12 animals received 100 mg methyl-prednisolone (orally) daily for 3 months. Their 12 sister pigs served as controls and received no corticosteroid. After sacrificing the animal, resistance arteries (diameter approximately 250 μm) were isolated from the femoral head epiphyseal cancellous bone and mounted as ring preparations on a small vessel myograph for measurement of isometric force development. The vasocontractory response to increasing doses of noradrenaline was not altered by methylprednisolone treatment. After submaximal precontraction by noradrenaline, vasorelaxation by sodium nitroprusside was not altered by methylprednisolone treatment. Only part of both experimental groups showed vasorelaxation to substance P and increasing doses of bradykinin. Increasing dose of endothelin-1 evoked vasoconstriction in both experimental groups. Sensitivity to endothelin-1 was increased after 3 months of methylprednisolone treatment. The response of isolated intraosseous femoral head arteries to endothelin-1 after long term glucocorticoid treatment in the pig is enhanced and appears to be unchanged for other physiologic vasoactive substances.

Short-term Preservation of Porcine Oocytes in Ambient Temperature: Novel Approaches

The objective of this study was to evaluate the feasibility of preserving porcine oocytes without freezing. To optimize preservation conditions, porcine cumulus-oocyte complexes (COCs) were preserved in TCM-199, porcine follicular fluid (pFF) and FCS at different temperatures (4°C, 20°C, 25°C, 27.5°C, 30°C and 38.5°C) for 1 day, 2 days or 3 days. After preservation, oocyte morphology, germinal vesicle (GV) rate, actin cytoskeleton organization, cortical granule distribution, mitochondrial translocation and intracellular glutathione level were evaluated. Oocyte maturation was indicated by first polar body emission and spindle morphology after in vitro culture. Strikingly, when COCs were stored at 27.5°C for 3 days in pFF or FCS, more than 60% oocytes were still arrested at the GV stage and more than 50% oocytes matured into MII stages after culture. Almost 80% oocytes showed normal actin organization and cortical granule relocation to the cortex, and approximately 50% oocytes showed diffused mitochondria distribution patterns and normal spindle configurations. While stored in TCM-199, all these criteria decreased significantly. Glutathione (GSH) level in the pFF or FCS group was higher than in the TCM-199 group, but lower than in the non-preserved control group. The preserved oocytes could be fertilized and developed to blastocysts (about 10%) with normal cell number, which is clear evidence for their retaining the developmental potentiality after 3d preservation. Thus, we have developed a simple method for preserving immature pig oocytes at an ambient temperature for several days without evident damage of cytoplasm and keeping oocyte developmental competence.

Effects of acute fasting and age on leptin and peroxisome proliferator-activated receptor gamma production relative to fat depot in immature and mature pigs

Leptin and peroxisome proliferator-activated receptor gamma (PPARγ) are adipogenic proteins that are actively involved in metabolic homeostasis of fat. Recently, it was reported that fat tissue in humans and rodents differs in metabolic activity relative to anatomical location of the fat tissue (i.e. depots) and animal age. Hence, we hypothesized that leptin and PPARγ production in various fat depots in female pigs differs in response to acute fasting, and that these responses vary with physiological maturity of the animal. Sixteen intact crossbred immature female pigs [prepubertal (PP); 132.2 ± 4.1 days] and 16 sexually mature female pigs (M; 224 ± 7.4 days) housed in an open-air, concrete slab, sheltered barn were randomly assigned to either Control or Fasted treatments. Control pigs (PP, n = 8; M, n = 8) had ad libitum access to feed, while Fasted pigs (PP, n = 8; M, n = 8) were denied access to feed from the onset of the study (0 h) to euthanasia at 72 h. Immediately post-mortem, fat samples were collected from the subcutaneous, pelvic, kidney, and heart (M pigs only) fat depots and analysed for leptin and PPARγ mRNA and protein content. Acute fasting decreased mean leptin mRNA tissue content in a depot specific manner in M pigs (p < 0.01), while mean leptin protein concentrations in fat tissues did not differ with fat depot or age of the pig. Furthermore, acute fasting did not affect mean PPARγ mRNA tissue content in a fat depot or age dependent manner. Mean concentrations of PPARγ protein in fat depots tended to be greater in M vs. PP pigs (p = 0.07). We suggest that these data provide evidence that acute fasting has a greater effect on leptin than PPARγ production in a fat depot dependent manner in M pigs, which may be indicative of changing physiological demands as an animal matures.

Neurocentral Synchondrosis Screws to Create and Correct Experimental Deformity: A Pilot Study

Unilateral pedicle screw epiphysiodesis of the neurocentral synchondrosis (NCS) can produce asymmetric growth of the synchondrosis to create scoliosis in an immature animal model. We asked whether a preexisting experimentally created scoliosis could be limited and corrected by modulating the growth of the faster-growing NCS by a similar method. Nine 1-month-old pigs were assigned to each of three groups: (1) a sham group in which three animals received a sham operation but without a pedicle screw fixation; (2) an experimental group with double right pedicle screws placed across the NCS from T7 to T14 (scoliosis-untreated); and (3) an experimental group treated in the same way except a second set of double pedicle screws was placed in the left pedicles 6weeks after the screws were placed on the right (scoliosis-treated). All animals were euthanized at 17weeks, and radiographs and axial CT images of the spine were obtained. A scoliotic curve was not seen in any of the animals in the sham group, in three of three in the scoliosis-untreated group with an average of 34°, and in three of three in the scoliosis-treated group with an average of 20°. In comparison to the scoliosis-untreated group, the second set of pedicle screws produced a 41% correction of the scoliosis. We found the pedicle screw inhibited the overgrowth of the NCS to prevent further curve progression and obtained some correction of the deformity. The NCS screw epiphysiodesis can create and reverse scoliosis in an immature pig model.

Correlations between effective permeability and marrow contact channels surface of vertebral endplates

Homeostasis of the intervertebral disc relies on nutrient supply and waste clearance through the dense capillary network that is in contact with the cartilage endplate (CEP). We developed a micro-computerized tomography (micro-CT) method to quantify the marrow contact channel surface (MCCS) with the CEP and to validate the hypothesis according to which MCCS was correlated to the effective permeability of the vertebral endplate (VEP) and influenced by the mechanical stimuli. The influence of compression loading on local vascularization was investigated. Six 4-week-old skeletally immature pigs were instrumented with left pedicle screws and rod at both T5-T6 and L1-L2 levels to create asymmetrical spine tethers. After 3 months of growth, three cylindrical specimens of the VEP (one central and two lateral right and left) were obtained from both the instrumented and the control levels. We used a previously validated method for measuring permeability. Micro-CT analysis (resolution 12 microm) yielded a gray-scale 2D-image of the discal end of each specimen converted into a binary 2D-image to derive the MCCS. Correlations between MCCS and effective permeability were assessed. Effective permeability and MCCS were significantly decreased compared to the control group especially on the tethered side (-41.5%, p = 0.004 and -52.5%, p = 0.0009, respectively). Correlations were significant and showed maximal value (r(2) = 0.430, p < 0.0001) on the tethered side involving maximal compressive loadings. Mechanical stimuli, due to unbalanced growth, altered the vascularization and the convective properties of the CEP. The cascade of mechanobiological events should offer perspectives for research on disc degeneration and attempted treatment.

Immunohistochemical studies on the proliferative marker Ki-67 and estrogen receptor alpha (ERα) in the uterus of neonatal and immature pigs following exposure to flutamide

The development of uterine glands is characterized by the proliferation of epithelial cells and by estrogen receptor alpha (ERα) expression in the nascent glandular epithelium. It is known that androgen receptors are present in the porcine uterus during prenatal development and the neonatal window, when adenogenesis occurs. Therefore, the objective of the study was to determine whether the effects of maternal or neonatal administration of the anti-androgen, flutamide, could entail changes in the presence of ERα and proliferation of uterine cells in neonatal and three-month-old pigs. Following prenatal flutamide exposure, morphological differences and the acceleration of uterus differentiation marked by ERα expression in epithelial crypts were observed in the neonatal piglets. In the three-month-old pig uterus, the proliferation of stromal cells was observed only after prenatal exposure to flutamide, whereas ERα staining was weaker. The neonatal administration of flutamide caused a significant decrease in the proliferation of the surface epithelium and diminished intensity of ERα staining in the stromal cells of the uterus of three-month-old pigs, which paralleled decreased estrogen levels in these animals. Overall, prenatal flutamide exposure promoted growth and development of the neonatal porcine uterus. Moreover, in three-month-old pigs, flutamide application during the neonatal period decreased surface epithelium proliferation and stromal ERα expression, which confirmed the importance of epithelial–stromal interactions in the adenogenesis.

The Effect of Posterior Distraction on Vertebral Growth in Immature Pigs

Experimental study. The aim of this study is to evaluate the vertebral body growth under distraction forces in immature pigs treated with growing rod (GR) technique. Distraction forces applied on growth plate of appendicular skeleton stimulate longitudinal growth. However, the effect of distraction forces on axial skeletal growth has not been fully investigated yet. Twelve 10-week-old domestic pigs were used in this experimental model to simulate GR technique. Four of them were lost during postoperative period because of deep wound infection. Cranially T12-L1 and caudally L4-L5 vertebrae were instrumented by pedicle screws bilaterally, while L2 and L3 were skipped. Distraction between pedicle screws was applied at index surgery. The rods were then lengthened twice in a month interval. All subjects were evaluated with anteroposterior and lateral spinal radiograph before surgery, after surgery, and at the final follow-up. The vertebral body heights of distracted segments (HD = L2 and L3) and control segments (HC = T9, T10 and T11) were measured. Average vertebral body heights and the increase percentage in the vertebral body heights were compared among control segments and distracted segments. The preoperative vertebral body height was similar in 2 groups (preHC: 10.81 mm, n = 19, preHD: 11.27 mm, n = 16, P > 0.05). At the final follow-up, the average vertebral body height in distraction group was significantly higher than the control group (postHC: 17.03 mm, postHD: 18.58 mm, P < 0.05). The increase percentage in vertebral body height was higher in distracted segments, but there was no statistically significant difference between the 2 groups. The vertebral growth continues during GR instrumentation. Distraction forces might stimulate also apophyseal growth of axial skeleton.

Expression pattern of acetylated α‐tubulin in porcine spermatogonia

Mammalian spermatogonial stem cells reside on the basement membrane of the seminiferous tubules. The mechanisms responsible for maintenance of spermatogonia at the basement membrane are unclear. Since acetylated alpha-tubulin (Ac-alpha-Tu) is a component of long-lived, stable microtubules and deacetylation of alpha-tubulin enhances cell motility, we hypothesized that acetylation of alpha-tubulin might be associated with positioning of spermatogonia at the basement membrane. The expression pattern of Ac-alpha-Tu at different stages of testis development was characterized by immunohistochemistry for Ac-alpha-Tu and spermatogonia-specific proteins (PGP 9.5, DAZL). In immature pig testes, Ac-alpha-Tu was present exclusively in gonocytes at 1 week of age, and in a subset of spermatogonia at 10 weeks of age. At this age, spermatogonia are migrating toward the tubule periphery and Ac-alpha-Tu appeared polarized toward the basement membrane. In adult pig testes, Ac-alpha-Tu was detected in few single or paired spermatogonia at the basement membrane as well as in spermatids and spermatozoa. Only undifferentiated (DAZL-), proliferating (determined by BrdU incorporation) spermatogonia expressed high levels of Ac-alpha-Tu. Comparison with the expression pattern of beta-tubulin and tyrosinated alpha-tubulin confirmed that only Ac-alpha-Tu is specific to germ cells. The unique pattern of Ac-alpha-Tu in undifferentiated germ cells during postnatal development suggests that posttranslational modifications of microtubules may play an important role in recruiting and anchoring spermatogonia at the basement membrane. Mol. Reprod. Dev. 77: 348-352, 2010. (c) 2009 Wiley-Liss, Inc.

Collagen-Platelet Composite Enhances Biomechanical and Histologic Healing of the Porcine Anterior Cruciate Ligament

Background The anterior cruciate ligament (ACL) fails to heal after traumatic rupture. Furthermore, large-animal models have recently shown that 1-month functional ACL healing is augmented after suture repair when a bioactive scaffold is placed in the tear site. Hypothesis At the time of suture repair, placement of a bioactive scaffold in the ACL wound site would improve the structural properties of the tissue. Study Design Controlled laboratory study. Methods Twenty-seven knees in immature pigs underwent ACL transection and suture repair. A collagen-platelet composite (CPC) was used to supplement the repair in 14 knees. Knees were harvested at 4 weeks, 6 weeks, and 3 months. Mechanical testing and histologic analysis were performed. Results The addition of a CPC to a suture repair resulted in improvements in yield load and linear stiffness of the repair tissue at 3 months, as well as a significant increase in cell density. A reduction in yield load and stiffness occurred at the 6-week time point in both groups, a phase when revascularization was noted. Conclusion The addition of a CPC to a suture repair enhanced the structural properties of the ACL, and the improvement was associated with increased cellularity within the healing ligament. Clinical Relevance The addition of a bioactive scaffold to the wound site improved the functional healing of the ACL after suture repair. The decreased repair strength during revascularization may indicate a need to protect the repair site through this period.

Influence of asymmetric tether on the macroscopic permeability of the vertebral end plate

We implemented an experimental model of asymmetrical compression loading of the vertebral end plate (VEP) in vivo. The macroscopic permeability of the VEP was measured. We hypothesized that static asymmetrical loading on vertebrae altered the macroscopic permeability of the VEP. In scoliosis, solute transport to and from the disc is dramatically decreased especially at the apical intervertebral disc. The decrease in permeability could be induced by mechanical stress. Nine skeletally immature pigs were instrumented with left pedicle screws and compression rod at the T5/T6 and L1/L2 levels. After 3 months, three cylindrical specimens of the VEP were obtained from each of the tethered levels. A previously validated method for measuring permeability, based on the relaxation pressure due to a transient-flow rate was used. A pistoning device generated a fluid flow that fully saturated the cylindrical specimen. The decrease in upstream pressure was measured using a pressure transducer, which allowed the macroscopic permeability to be derived. A microscopic study completed the approach. Overall macroscopic permeability was lower for the tethered VEPs than for the VEPs of the control group, respectively -47% for flow-in (p = 0.0001) and -46% for flow-out (p = 0.0001). In the tethered group, macroscopic permeability of the specimens from the tethered side was lower than macroscopic permeability of those from the non-tethered side, -39% for flow-out (p = 0.024) and -47% for flow-in (p = 0.13). In the control group, the macroscopic permeability was greater in the center of the VEP than in its lateral parts for flow-out (p = 0.004). Macroscopic permeability of the center of the VEPs was greater for flow-out than for flow-in (p = 0.02). There was no significant difference between thoracic and lumbar. This study demonstrated that compression loading applied to a growing spine results in decreased permeability of the VEP. This result could be explained by local remodeling, such as calcification of the cartilage end plate or sclerosis of the underlying bone.

Expression and activities of 11βHSD enzymes in the testes and reproductive tracts of sexually immature male pigs

In light of studies implicating glucocorticoids in the control of testicular steroidogenesis and/or spermatogenesis, the objective of this study was to characterise the expression and activities of the 11β-hydroxysteroid dehydrogenase (11βHSD) enzymes in the testis and reproductive tract of the pre-pubertal pig. Although 11βHSD1 and 11βHSD2 mRNA transcripts and proteins were co-expressed in all regions of the reproductive tract, cortisol–cortisone inter-conversion was detectable in the testis, caput epididymidis and bulbourethral glands only. In homogenates of these 3 tissues, the apparent K m for NADP +- and NAD +-dependent 11β-dehydrogenase activities ranged between 152–883 and 47–479 nmol l −1, respectively. Irrespective of the pyridine nucleotide co-substrate, estimates of V max were consistently two orders of magnitude higher in the testis. Moreover, while, in each tissue, levels of cortisol oxidation were comparable in the presence of either NADP + or NAD +, maximal rates of NAD(P) +-dependent cortisol oxidation were up to 33-fold greater than the V max for NADPH-dependent reduction of cortisone. We conclude that in the testis, caput epididymidis and bulbourethral gland of the immature pig, NADP +- and NAD +-dependent 11βHSD enzymes catalyse net inactivation of cortisol, suggesting a physiological role for these enzymes in limiting local actions of glucocorticoids within these male reproductive tissues prior to puberty.

Hypoxia and HIF-1α expression in the epiphyseal cartilage following ischemic injury to the immature femoral head

HIF-1α has been shown to be a central mediator of cellular response to hypoxia. The role it plays after ischemic injury to the immature femoral head is unknown. The purpose of this study was to determine the region of the femoral head affected by hypoxia following ischemic injury to the immature femoral head and to determine the site of HIF-1α activation and revascularization. We hypothesize that the epiphyseal cartilage, rather than the bony epiphysis, is the site of HIF-1α activation following ischemic osteonecrosis and that the epiphyseal cartilage plays an important role in the revascularization process. Materials and methodsFemoral head osteonecrosis was surgically induced in 56 immature pigs. Hypoxyprobe staining, cell viability assay, HIF-1α western blot, RT-qPCR of HIF-1α, VEGF, VEGFR2, and PECAM, and micro-CT assessments of microfil-infused femoral heads were performed. ResultsSevere hypoxia was present in the bony epiphysis and the lower part of the epiphyseal cartilage following ischemia. In the bony epiphysis, extensive cell death and tissue necrosis was observed with degradation of proteins and RNAs which precluded further analysis. In the epiphyseal cartilage, the loss of cell viability was limited to its deep layer with the remainder of the cartilage remaining viable. Furthermore, the cartilage from the ischemic side showed a significant increase in HIF-1α protein level and HIF-1α expression. VEGF expression in the cartilage was dramatically and significantly increased at 24 h, 2 and 4 weeks (p<0.05 for all) with 5 to 10 fold increase being observed on the ischemic side compared to the normal side. PECAM and VEGFR2 expressions in the cartilage were both significantly decreased at 24 h but returned to the normal levels by 2 and 4 weeks, respectively. Micro-CT showed revascularization of the cartilage on the ischemic side with the vessel volume/total volume equaling the normal side by 4 weeks. ConclusionsAcute ischemic injury to the immature femoral head induced severe hypoxia and cell death in the bony epiphysis and the deep layer of the epiphyseal cartilage. Viable chondrocytes in the superficial layer of the epiphyseal cartilage showed HIF-1α activation and VEGF upregulation with subsequent revascularization occurring in the cartilage.

Collagen-Platelet Composites Improve the Biomechanical Properties of Healing Anterior Cruciate Ligament Grafts in a Porcine Model

Background The outcome of anterior cruciate ligament (ACL) reconstruction is variable, and many patients have increased joint laxity postoperatively. Hypothesis Placement of a collagen-platelet composite (CPC) around the graft at the time of ACL reconstruction decreases postoperative knee laxity and improves the structural properties of the graft compared with standard ACL reconstruction. Study Design Controlled laboratory study. Methods Thirteen immature pigs underwent unilateral ACL reconstruction with a bone–patellar tendon–bone allograft. In 6 pigs, a standard allograft was used to reconstruct the ACL. In 7 pigs, a CPC was placed around the allograft. After 15 weeks of healing, the animals were euthanized, and the anterior-posterior (AP) knee laxity and structural properties of the graft were measured. Qualitative histology of the grafts was also performed. Results The AP laxity values of the reconstructed knees, normalized to the contralateral control, were significantly reduced by 28% and 57% at 60° and 90° of knee flexion, respectively, with the addition of CPC (P <. 001). Significant improvements in the graft structural properties were also found; the normalized yield (P =. 044) and maximum failure loads (P =. 025) of the CPC group were 60% higher than the standard ACL-reconstructed group. Although cellular and vessel infiltration were observed in the grafts of both groups, regions of necrosis were present only in the standard ACL-reconstructed group. Conclusion These data demonstrate that the application of CPC at the time of ACL reconstruction improves the structural properties of the graft and reduces early AP knee laxity in the porcine model after 15 weeks of healing. Clinical Relevance Application of a CPC to an ACL graft at the time of surgery decreased knee laxity and increased the structural properties of the graft after 15 weeks of healing.

Leptin and Its Receptor Are Expressed in the Testis and in the Epididymis of Young and Adult Pigs

Recent studies indicated that leptin, a 16 kDa hormone, is a regulatory signal in human and rodent male reproduction. This work was designed to investigate the expression of leptin and its receptor in testes and epididymides from immature and mature pigs. Immunolocalization revealed that leptin and its receptor were confined only in the interstitial compartment of immature testes, whereas both proteins were detected in Leydig cells and within seminiferous tubules of mature gonads. The immunostaining of epididymal tissues showed that leptin was absent in the epithelial cells of immature pigs but it was present in all the three regions of mature epididymides, although with a minor signal in the cauda. Conversely, leptin receptor was observed in all the epithelial cells of both immature and mature epididymides. Western blot analysis of tissue extracts detected a 16 kDa band for leptin and five/six isoforms, ranging from 120 to 40 kDa, for leptin receptor. In conclusion, this work has identified, for the first time, leptin and leptin receptor in the testis and in the epididymis of the pig showing a differential cell-type expression pattern of the two proteins in young and adult animals. Therefore, our findings suggest a possible involvement of leptin in endocrine or autocrine/paracrine control of porcine male reproductive structures.

Spinal Hemiepiphysiodesis Decreases the Size of Vertebral Growth Plate Hypertrophic Zone and Cells

Hemiepiphysiodesis is a potential method to treat idiopathic juvenile scoliosis early. The purpose of the present study was to investigate a mechanism of curve creation in the pig thoracic model of spinal hemiepiphysiodesis by determining whether the structure of the vertebral growth plate varied with distance from the stapled, concave side of the spine. The hypotheses were that the heights of the hypertrophic zone, hypertrophic cells, and disc would be decreased on the treated side of the treated level as compared with both an unstapled control level and the side opposite the staple. Custom spine staples were implanted into six midthoracic vertebrae in each of five skeletally immature pigs. After eight weeks, the spines were harvested and histological sections were prepared. Hypertrophic zone height, hypertrophic cell height and width, and disc height were measured at discrete coronal plane locations at stapled and unstapled thoracic levels. Differences between stapled and unstapled levels and locations were compared with use of mixed linear modeling for repeated measures, followed by regression models to determine growth plate intercept and slope across the plane by thoracic level. Zone height, cell height, and cell width were lowest on the stapled side of the stapled level, with significant differences in the overall statistical model (p < 0.02). Disc heights were significantly reduced (p < 0.0001) at the stapled levels across the coronal plane. Unilateral control of intervertebral joint motion decreased growth plate height, cell size, and disc height.

Extracorporeal cell therapy with granulocytes in a pig model of Gram-positive sepsis

Granulocyte transfusions have been used to treat immune cell dysfunction in sepsis. As granulocyte transfusions can trigger tissue injury via local effects of neutrophils, we hypothesized that extracorporeal treatment of plasma using granulocytes would prove beneficial while having less side effects. Prospective controlled three-armed animal study. Research laboratory. Twenty-one female immature pigs (7.5-12 kg, 7-9 weeks old). Three groups of spontaneously breathing, sedated pigs (n = 7 each) received an intravenous lethal dose of live Staphylococcus aureus over 1 hour. Although group I had no specific treatment (control), group II and III were subsequently treated for 4 hours with an extracorporeal device containing either no cells (sham control, group II) or human cell line-derived granulocytic cells (group III). Survival time and physiologic, biochemical, and hematologic parameters were monitored for 7 days. All animals of group I died during the observation time (mean survival time: 70 hours). In group II, two of seven and in group III, six of seven animals survived the observation time (mean survival: 75 and 168 hours, respectively). Survival differences were significant between group I and III (p < 0.001) and between group II and III (p < 0.05) but not between group I and II (p = 0.43). Furthermore, group differences in bacterial blood concentrations, differential blood count, blood gases, lactate, and interleukins were observed. The extracorporeal cell treatment was well tolerated by the animals. Extracorporeal therapy with granulocytic cells significantly improved survival in a pig model of sepsis. Further studies with this approach are encouraged.