E8002 Reduces Adhesion Formation and Improves Joint Mobility in a Rat Model of Knee Arthrofibrosis.

Knee arthrofibrosis is a common complication of knee surgery, caused by excessive scar tissue, which results in functional disability. However, no curative treatment has been established. E8002 is an anti-adhesion material that contains L-ascorbic acid, an antioxidant. We aimed to evaluate the efficacy of E8002 for the prevention of knee arthrofibrosis in a rat model, comprising injury to the surface of the femur and quadriceps muscle 1 cm proximal to the patella. Sixteen male, 8-week-old Sprague Dawley rats were studied: in the Adhesion group, haemorrhagic injury was induced to the quadriceps and bone, and in the E8002 group, an adhesion-preventing film was implanted between the quadriceps and femur after injury. Six weeks following injury, the restriction of knee flexion owing to fibrotic scarring had not worsened in the E8002 group but had worsened in the Adhesion group. The area of fibrotic scarring was smaller in the E8002 group than in the Adhesion group (p < 0.05). In addition, the numbers of fibroblasts (p < 0.05) and myofibroblasts (p < 0.01) in the fibrotic scar were lower in the E8002 group. Thus, E8002 reduces myofibroblast proliferation and fibrotic scar formation and improves the range of motion of the joint in a model of knee injury.

E8002 Inhibits Peripheral Nerve Adhesion by Enhancing Fibrinolysis of l-Ascorbic Acid in a Rat Sciatic Nerve Model.

Perineural adhesions leading to neuropathy are one of the most undesirable consequences of peripheral nerve surgery. However, there are currently no widely used compounds with anti-adhesive effects in the field of peripheral nerve surgery. E8002 is a novel, anti-adhesive, multi-layer membrane that contains L-ascorbic acid (AA). Here, we investigated the effect and mechanism of E8002 in a rat sciatic nerve adhesion model. A total of 21 rats were used. Six weeks after surgery, macroscopic adhesion scores were significantly lower in the E8002 group (adhesion procedure followed by nerve wrapping with E8002) compared to the E8002 AA(-) group (adhesion procedure followed by nerve wrapping with the E8002 membrane excluding AA) and adhesion group (adhesion procedure but no treatment). Correspondingly, a microscopic examination revealed prominent scar tissue in the E8002 AA(-) and adhesion groups. Furthermore, an in vitro study using human blood samples showed that AA enhanced tissue-type, plasminogen activator-mediated fibrinolysis. Altogether, these results suggest that E8002 may exert an anti-adhesive action via AA and the regulation of fibrinolysis.

Application of a Novel Anti-Adhesive Membrane, E8002, in a Rat Laminectomy Model.

Neuropathic pain after spinal surgery, so-called failed back surgery syndrome, is a frequently observed common complication. One cause of the pain is scar tissue formation, observed as post-surgical epidural adhesions. These adhesions may compress surrounding spinal nerves, resulting in pain, even after successful spinal surgery. E8002 is an anti-adhesive membrane. In Japan, a clinical trial of E8002 is currently ongoing in patients undergoing abdominal surgery. However, animal experiments have not been performed for E8002 in spinal surgery. We assessed the anti-adhesive effect of E8002 in a rat laminectomy model. The dura matter was covered with an E8002 membrane or left uncovered as a control. Neurological evaluations and histopathological findings were compared at six weeks postoperatively. Histopathological analyses were performed by hematoxylin⁻eosin and aldehyde fuchsin-Masson Goldner staining. Three assessment areas were selected at the middle and margins of the laminectomy sites, and the numbers of fibroblasts and inflammatory cells were counted. Blinded histopathological evaluation revealed that adhesions and scar formation were reduced in the E8002 group compared with the control group. The E8002 group had significantly lower numbers of fibroblasts and inflammatory cells than the control group. The present results indicate that E8002 can prevent epidural scar adhesions after laminectomy.

Host conditioning and rejection monitoring in hepatocyte transplantation in humans.

BACKGROUND & AIMS: Hepatocyte transplantation partially corrects genetic disorders and has been associated anecdotally with reversal of acute liver failure. Monitoring for graft function and rejection has been difficult, and has contributed to limited graft survival. Here we aimed to use preparative liver-directed radiation therapy, and continuous monitoring for possible rejection in an attempt to overcome these limitations. METHODS: Preparative hepatic irradiation was examined in non-human primates as a strategy to improve engraftment of donor hepatocytes, and was then applied in human subjects. T cell immune monitoring was also examined in human subjects to assess adequacy of immunosuppression. RESULTS: Porcine hepatocyte transplants engrafted and expanded to comprise up to 15% of irradiated segments in immunosuppressed monkeys preconditioned with 10Gy liver-directed irradiation. Two patients with urea cycle deficiencies had early graft loss following hepatocyte transplantation; retrospective immune monitoring suggested the need for additional immunosuppression. Preparative radiation, anti-lymphocyte induction, and frequent immune monitoring were instituted for hepatocyte transplantation in a 27year old female with classical phenylketonuria. Post-transplant liver biopsies demonstrated multiple small clusters of transplanted cells, multiple mitoses, and Ki67+ hepatocytes. Mean peripheral blood phenylalanine (PHE) level fell from pre-transplant levels of 1343±48µM (normal 30-119µM) to 854±25µM (treatment goal ≤360µM) after transplant (36% decrease; p<0.0001), despite transplantation of only half the target number of donor hepatocytes. PHE levels remained below 900µM during supervised follow-up, but graft loss occurred after follow-up became inconsistent. CONCLUSIONS: Radiation preconditioning and serial rejection risk assessment may produce better engraftment and long-term survival of transplanted hepatocytes. Hepatocyte xenografts engraft for a period of months in non-human primates and may provide effective therapy for patients with acute liver failure. LAY SUMMARY: Hepatocyte transplantation can potentially be used to treat genetic liver disorders but its application in clinical practice has been impeded by inefficient hepatocyte engraftment and the inability to monitor rejection of transplanted liver cells. In this study, we first show in non-human primates that pretreatment of the host liver with radiation improves the engraftment of transplanted liver cells. We then used this knowledge in a series of clinical hepatocyte transplants in patients with genetic liver disorders to show that radiation pretreatment and rejection risk monitoring are safe and, if optimized, could improve engraftment and long-term survival of transplanted hepatocytes in patients.

1,5-anhydro-D-fructose induces anti-aging effects on aging-associated brain diseases by increasing 5'-adenosine monophosphate-activated protein kinase activity via the peroxisome proliferator-activated receptor-γ co-activator-1α/brain-derived neurotrophic factor pathway.

5'-Adenosine monophosphate-activated protein kinase (AMPK) is a metabolic sensor that serves as a cellular housekeeper; it also controls energy homeostasis and stress resistance. Thus, correct regulation of this factor can enhance health and survival. AMPK signaling may have a critical role in aging-associated brain diseases. Some in vitro studies have shown that 1,5-anhydro-D-fructose (1,5-AF) induces AMPK activation. In the present study, we experimentally evaluated the effects of 1,5-AF on aging-associated brain diseases in vivo using an animal model of acute ischemic stroke (AIS), stroke-prone spontaneously hypertensive rats (SHRSPs), and the spontaneous senescence-accelerated mouse-prone 8 (SAMP8) model. In the AIS model, intraperitoneal injection of 1,5-AF reduced cerebral infarct volume, neurological deficits, and mortality. In SHRSPs, oral administration of 1,5-AF reduced blood pressure and prolonged survival. In the SAMP8 model, oral administration of 1,5-AF alleviated aging-related decline in motor cognitive function. Although aging reduced the expression levels of peroxisome proliferator-activated receptor-γ co-activator-1α (PGC-1α) and brain-derived neurotrophic factor (BDNF), we found that 1,5-AF activated AMPK, which led to upregulation of the PGC-1α/BDNF pathway. Our results suggest that 1,5-AF can induce endogenous neurovascular protection, potentially preventing aging-associated brain diseases. Clinical studies are needed to determine whether 1,5-AF can prevent aging-associated brain diseases.

Disrupted regulation of ghrelin production under antihypertensive treatment in spontaneously hypertensive rats.

BACKGROUND: Ghrelin is an acylated peptide hormone mainly secreted from the stomach. When administrated externally it modulates vascular tone mainly through the regulation of autonomic nerve activity. However, the effects of blood pressure (BP) on the production and secretion of ghrelin remain to be clarified. METHODS AND RESULTS: We examined the stomach and plasma levels of ghrelin in spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats after a 4-week-intervention with antihypertensive agents (candesartan-cilexetil [ARB], doxazosin [DZN], metoprolol [MP], reserpine [RES]) to clarify the influence of BP on the secretion of ghrelin. The effect of these agents on ghrelin production and secretion were examined by comparing vehicle-treated controls (WKY-Intact, SHR-Intact). Treatment with the 4 antihypertensive drugs all yielded a significant decline in systolic BP in both SHR and WKY. Under these conditions, significantly lower levels of stomach and plasma ghrelin were detected in WKY treated with ARB (P<0.05), DZN (P<0.05), MP (P<0.05) and RES (P<0.05) compared with WKY-Intact, whereas no significant change in the ghrelin levels in the stomach and plasma were detected in SHR under the same treatments. CONCLUSIONS: The findings imply that the production and secretion of ghrelin are controlled by the ambient vascular tone and vice versa in normotensive WKY. This inter-relationship between ghrelin and BP seems to be disrupted in SHR.

Preconditioning Exercise in Rats Attenuates Early Brain Injury Resulting from Subarachnoid Hemorrhage by Reducing Oxidative Stress, Inflammation, and Neuronal Apoptosis.

Subarachnoid hemorrhage (SAH) is a catastrophic form of stroke responsible for significant morbidity and mortality. Oxidative stress, inflammation, and neuronal apoptosis are important in the pathogenesis of early brain injury (EBI) following SAH. Preconditioning exercise confers neuroprotective effects, mitigating EBI; however, the basis for such protection is unknown. We investigated the effects of preconditioning exercise on brain damage and sensorimotor function after SAH. Male rats were assigned to either a sham-operated (Sham) group, exercise (Ex) group, or no-exercise (No-Ex) group. After a 3-week exercise program, they underwent SAH by endovascular perforation. Consciousness level, neurological score, and sensorimotor function were studied. The expression of nuclear factor erythroid 2 p45-related factor 2 (Nrf2), heme oxygenase 1 (HO-1), 4-hydroxynonenal (4HNE), nitrotyrosine (NT), ionized calcium-binding adaptor molecule 1 (Iba1), tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), interleukin 1ß (IL-1ß), 14-3-3γ, p-ß-catenin Ser37, Bax, and caspase-3 were evaluated by immunohistochemistry or western blotting. The terminal deoxynucleotidyl transferase-mediated biotinylated dUTP nick end labeling (TUNEL) assay was also performed. After SAH, the Ex group had significantly reduced neurological deficits, sensorimotor dysfunction, and consciousness disorder compared with the No-Ex group. Nrf2, HO-1, and 14-3-3γ were significantly higher in the Ex group, while 4HNE, NT, Iba1, TNF-α, IL-6, IL-1ß, Bax, caspase-3, and TUNEL-positive cells were significantly lower. Our findings suggest that preconditioning exercise ameliorates EBI after SAH. The expression of 4HNE and NT was reduced by Nrf2/HO-1 pathway activation; additionally, both oxidative stress and inflammation were reduced. Furthermore, preconditioning exercise reduced apoptosis, likely via the 14-3-3γ/p-ß-catenin Ser37/Bax/caspase-3 pathway.

Uric acid enhances alteplase-mediated thrombolysis as an antioxidant.

Uric acid (UA) therapy may prevent early ischemic worsening after acute stroke in thrombolysis patients. The aim of this study was to examine the influence of UA on the thrombolytic efficacy of alteplase in human blood samples by measuring thrombolysis under flow conditions using a newly developed microchip-based flow-chamber assay. Human blood samples from healthy volunteers were exposed to UA, alteplase, or a combination of UA and alteplase. Whole blood and platelet-rich plasma were perfused over a collagen- and thromboplastin-coated microchip, and capillary occlusion was monitored with a video microscope and flow-pressure sensor. The area under the curve (extent of thrombogenesis or thrombolysis) at 30 minutes was 92% lower in the UA-alteplase-treated group compared with the alteplase-treated group. D-dimers were measured to evaluate these effects in human platelet-poor plasma samples. Although hydrogen peroxide significantly decreased the elevation of D-dimers by alteplase, UA significantly inhibited the effect of hydrogen peroxide. Meanwhile, rat models of thromboembolic cerebral ischemia were treated with either alteplase or UA-alteplase combination therapy. Compared with alteplase alone, the combination therapy reduced the infarct volume and inhibited haemorrhagic transformation. UA enhances alteplase-mediated thrombolysis, potentially by preventing oxidative stress, which inhibits fibrinolysis by alteplase in thrombi.

Characteristics and gender differences concerning pulmonary hemodynamics in Clawn miniature pigs.

The characteristics and gender differences of the pulmonary hemodynamic parameters of 16 Clawn miniature pigs were examined and the data were compared with reports concerning dogs and other pig species. The pulmonary systolic, diastolic and mean arterial blood pressures of the mini-pig were slightly higher than those of the dog, respectively, but both the right atrial pressure and pulmonary capillary wedge pressure were within the normal physiological ranges of the dog. Concerning gender differences in hemodynamic parameters of the mini-pig, the female values, except the right atrial pressure, were slightly higher than those of the male, but no significant differences were recognized. The present study results will help pulmonary researchers understand the differences between Clawn miniature pigs and dogs for accurate analysis of experimental results.

Skin morphology of the Clawn miniature pig.

Skin morphology of the Clawn miniature pig (CMP) was investigated at the axilla, medial thigh, back and loin. The mean thickness of the epidermis (excluding the corneal layer), the mean number of layers of keratinocytes comprising the epidermis and the mean height of keratinocytes were assessed morphometrically. When observed under a light microscope, the skin of the CMP resembled human skin. Morphometrically, skin from the back and loin of the CMP most resembles human skin. Electron microscopic observations revealed sparse but typical Birbeck granules in the epidermal Langerhans cells of the CMP. The results of the present study indicate that CMP skin is potentially useful as a model for human skin.

Edaravone, a Synthetic Free Radical Scavenger, Enhances Alteplase-Mediated Thrombolysis.

The combination of alteplase, a recombinant tissue plasminogen activator, and edaravone, an antioxidant, reportedly enhances recanalization after acute ischemic stroke. We examined the influence of edaravone on the thrombolytic efficacy of alteplase by measuring thrombolysis using a newly developed microchip-based flow-chamber assay. Rat models of embolic cerebral ischemia were treated with either alteplase or alteplase-edaravone combination therapy. The combination therapy significantly reduced the infarct volume and improved neurological deficits. Human blood samples from healthy volunteers were exposed to edaravone, alteplase, or a combination of alteplase and edaravone or hydrogen peroxide. Whole blood was perfused over a collagen- and thromboplastin-coated microchip; capillary occlusion was monitored with a video microscope and flow-pressure sensor. The area under the curve (extent of thrombogenesis or thrombolysis) at 30 minutes was 69.9% lower in the edaravone-alteplase- than alteplase-treated group. The thrombolytic effect of alteplase was significantly attenuated in the presence of hydrogen peroxide, suggesting that oxidative stress might hinder thrombolysis. D-dimers were measured to evaluate these effects in human platelet-poor plasma samples. Although hydrogen peroxide significantly decreased the elevation of D-dimers by alteplase, edaravone significantly inhibited the decrease. Edaravone enhances alteplase-mediated thrombolysis, likely by preventing oxidative stress, which inhibits fibrinolysis by alteplase in thrombi.

Effects of olprinone on myocardial ischemia-reperfusion injury in dogs.

We investigated the effect of olprinone on canine myocardial pump function and myocardial damage after ischemia-reperfusion injury. Three dogs of the experimental group were given olprinone (Olprinone group) and another 3 dogs were served as control (Intact group). All animals were occluded left anterior descending artery for 60 min, followed by 6 hr of reperfusion. In the experiment, hemodynamics, infarct area, creatine kinase and troponin-I were measured. Olprinone infusion induced significantly high cardiac output value and significantly low values in left ventricular end diastolic pressure and systemic vascular resistance index after reperfusion. Also, olprinone tend to attenuate the infarct area, creatine kinase and troponin-I.

Discrepancy between Doppler and catheter measurements of pressure gradients across small-size prosthetic valve.

OBJECTIVES: The exact role of pressure gradient across the prosthetic valve estimated from Doppler flow velocity remains controversial. This in-vivo study was designed to assess the actual discrepancy between Doppler and catheter measurements of the pressure gradients for small bileaflet prosthetic valves in the aortic position. METHODS: Bileaflet prosthetic valves (19 mm-ATS) were implanted into the aortic position in pigs, and pressure gradients across the valves were examined by volume loading under right heart bypass. The pressure gradient obtained by catheter was defined as the conventional peak-to-peak gradient between the left ventricle and aorta. The peak Doppler gradients were calculated from the maximal instantaneous Doppler velocity with the ultrasound probe positioned on the diaphragm at the level of the cardiac apex. RESULTS: There were strong correlations between pressure gradients and cardiac output. The Doppler gradient was constantly higher than the catheter values, and the resultant discrepancy between Doppler and catheter measurements was directly dependent on cardiac output (y=9.9x+0.6, r2=0.55). For cardiac output > or = 5.0 L/min, the difference between Doppler and catheter gradients reached 40 mmHg, and maximum differences of up to 80 mmHg were observed. CONCLUSIONS: In view of the presence of striking overestimation of catheter gradient by Doppler measurement, Doppler ultrasound should be used cautiously to assess small-size bileaflet prosthetic valve function with consideration of the patient's hemodynamic state.

Resetting the transcription factor network reverses terminal chronic hepatic failure.

The cause of organ failure is enigmatic for many degenerative diseases, including end-stage liver disease. Here, using a CCl4-induced rat model of irreversible and fatal hepatic failure, which also exhibits terminal changes in the extracellular matrix, we demonstrated that chronic injury stably reprograms the critical balance of transcription factors and that diseased and dedifferentiated cells can be returned to normal function by re-expression of critical transcription factors, a process similar to the type of reprogramming that induces somatic cells to become pluripotent or to change their cell lineage. Forced re-expression of the transcription factor HNF4α induced expression of the other hepatocyte-expressed transcription factors; restored functionality in terminally diseased hepatocytes isolated from CCl4-treated rats; and rapidly reversed fatal liver failure in CCl4-treated animals by restoring diseased hepatocytes rather than replacing them with new hepatocytes or stem cells. Together, the results of our study indicate that disruption of the transcription factor network and cellular dedifferentiation likely mediate terminal liver failure and suggest reinstatement of this network has therapeutic potential for correcting organ failure without cell replacement.

Effects of propofol-sevoflurane anesthesia on the maternal and fetal hemodynamics blood gases, and uterine activity in pregnant goats.

To determine the effects of propofol and sevoflurane on hemodynamics, acid-base balance and uterine activity in pregnant animals, a prospective experimental study was designed by use of ten pregnant goats. Propofol was intravenously administered at a bolus dose of 5 mg/kg and then infused a rate of 0.3 mg/kg/min for 5 min. Following the induction, the animals were incrementally inhaled 2.7 and 4.1% of end-tidal concentration of sevoflurane each for 30 min, and then recovered. The maternal and fetal heart rate (HR), arterial blood pressure (BP) and acid-base balance, the intrauterine pressure (IUP), and the uterine blood flow (UBF) were measured. Following the pre-anesthetic data, the parameters were measured 7 times throughout the anesthetic and recovering periods. The propofol infusion induced 1.37 times of HR increase and produced decrease in PO(2) and a relevant metabolic acidemia in the mother, with no effect in the fetus. Sevoflurane reduced BP in the fetus from 30 (2.7%) to 60 (4.1%) min of inhalation. The uterine contractions disappeared throughout sevoflurane inhalation, and then recurred within 15 min after the cessation of sevoflurane. Propofol injection increases HR, and induces a moderate hypoxemia and metabolic acidemia associated with the suppressed ventilation for pregnant goats, with less effect on the fetal hemodinamics. Sevoflurane causes minimal change in maternal hemodynamics, but induces significant hypotension in the fetus and reduction of uterine activity. These data may be useful in making anesthetic choices combined with analgesia for Caesarian section in goats.

Effects of milrinone on hemodynamics and regional blood flow in the hypoxic dog.

Milrinone, a therapeutic agent for acute congestive heart failure, has both inotropic and vasodilatory effects, but investigations of these effects of milrinone were almost all conducted under normoxia, and few reports have investigated how milrinone affects the hemodynamics and redistribution of regional blood flow under severe hypoxia. By using colored microspheres, we investigated how milrinone affects hemodynamics and the redistribution of regional blood flow under severe hypoxia. Twelve healthy mongrel dogs were divided into 2 groups. The milrinone group was infused with milrinone cumulatively at 25, 75 and 250 microg/kg for 5 min each. The intact group was infused with saline instead of milrinone. We measured the hemodynamics and cerebrum, cerebellum and kidney blood flow in both groups. Both groups were inspired with 10% oxygen. Milrinone induced significant decrease in mean pulmonary artery and pulmonary vascular resistance, compared with the intact group. In both groups slight decreases in mean arterial pressure, systemic vascular resistance and double-product were seen. In regional blood flow, milrinone-induced increases in blood flow were seen in the cerebrum, cerebellum, and especially in the kidneys. Milrinone's vasodilatory effects were sufficient even under hypoxia. And milrinone increased regional blood flow slightly in the cerebrum and cerebellum, and significantly in the kidneys. These results suggested that milrinone protects against hypoxia-induced organ damage especially in the kidneys. In addition, milrinone is very potent in improving severe congested hemodynamics which complicates hypoxic pulmonary vasoconstriction.

Successful use of an acceleration rate response pacemaker with a transvenous steroid-eluting screw-in lead for third-degree atrioventricular block in a labrador retriever.

Permanent pacemakers are commonly used in veterinary practice and can have a dramatic effect on the treatment of heart block. A Labrador Retriever dog suffering from exercise intolerance secondary to third degree atrioventricular block was treated with a new pacemaker system. A steroid-eluting screw-in type lead that has the advantage of being more fixed to the myocardial wall without increasing the pacing threshold was used. The heart rate was regulated with an acceleration sensing pacemaker generator that included several automatic modulation systems. Nineteen months after implantation, the dog has a normal level of activity. The present case suggests that this pacemaker design may offer important advantages for canine patients.

Protective effect of neutralization of the extracellular high-mobility group box 1 on renal ischemia-reperfusion injury in miniature swine.

BACKGROUND: Strategies that reduce ischemia-reperfusion injury (IRI) have the potential to expand the numbers of available organs for transplantation. Recent reports in rodent models have demonstrated that high-mobility group box 1 (HMGB1) acts as an alarm in initiating the inflammatory response resulting from ischemic injury. The aim of this study was to evaluate the cytoprotective effects of anti-HMGB1 antibodies on renal IRI in preclinical large animals. METHODS: One hundred twenty minutes of warm and 60 min of cold renal ischemia were induced in 8 CLAWN miniature swine. Three of eight animals received intravenous anti-HMGB1 antibody at 1 mg/kg just before the reperfusion of renal blood flow. Renal function was assessed by serum creatinine and renal biopsy. Serum levels of interleukin (IL)-1ß, IL-6, and HMGB1 were measured. RESULTS: The concentration of HMGB1 increased as early as 30 min after reperfusion and before the elevation of IL-1ß and IL-6. Serum creatinine levels were markedly elevated, peaking at a median of 5 days (peak creatinine levels: 11.6 ± 1.6 mg/dL) and recovering by day 14. Anti-HMGB1 antibody injection dramatically decreased renal damage as well as serum levels of HMGB1 associated with IRI. Renal function returned to near normal by day 9, and peak creatinine levels were markedly lower (7.4 ± 0.2 mg/dL), and biopsies possessed fewer pathologic changes when compared to the control group. CONCLUSION: In this study, we demonstrated the beneficial effects of perioperative administration of anti-HMGB1 antibody in reducing renal IRI in a clinically relevant, large animal model.

A nonhuman primate model of human radiation-induced venocclusive liver disease and hepatocyte injury.

BACKGROUND: Human liver has an unusual sensitivity to radiation that limits its use in cancer therapy or in preconditioning for hepatocyte transplantation. Because the characteristic veno-occlusive lesions of radiation-induced liver disease do not occur in rodents, there has been no experimental model to investigate the limits of safe radiation therapy or explore the pathogenesis of hepatic veno-occlusive disease. METHODS AND MATERIALS: We performed a dose-escalation study in a primate, the cynomolgus monkey, using hypofractionated stereotactic body radiotherapy in 13 animals. RESULTS: At doses ≥40 Gy, animals developed a systemic syndrome resembling human radiation-induced liver disease, consisting of decreased albumin, elevated alkaline phosphatase, loss of appetite, ascites, and normal bilirubin. Higher radiation doses were lethal, causing severe disease that required euthanasia approximately 10 weeks after radiation. Even at lower doses in which radiation-induced liver disease was mild or nonexistent, latent and significant injury to hepatocytes was demonstrated by asialoglycoprotein-mediated functional imaging. These monkeys developed hepatic failure with encephalopathy when they received parenteral nutrition containing high concentrations of glucose. Histologically, livers showed central obstruction via an unusual intimal swelling that progressed to central fibrosis. CONCLUSIONS: The cynomolgus monkey, as the first animal model of human veno-occlusive radiation-induced liver disease, provides a resource for characterizing the early changes and pathogenesis of venocclusion, for establishing nonlethal therapeutic dosages, and for examining experimental therapies to minimize radiation injury.

Hepatocyte growth factor sustains T regulatory cells and prolongs the survival of kidney allografts in major histocompatibility complex-inbred CLAWN-miniature swine.

BACKGROUND: Although 12 days of high dose of FK506 permits the induction of tolerance of fully major histocompatibility complex (MHC)-mismatched allogeneic kidneys in MGH-miniature swine, we found that the same dose of FK506 is insufficient to induce such tolerance CLAWN-miniature swine. The CLAWN swine model was therefore chosen to study the potential immunoregulatory effects of human-recombinant hepatocyte growth factor (HGF). METHODS: Ten CLAWN miniature swine received fully MHC-mismatched kidneys with 12 days (days 0-11) of FK506. Among these 10 recipients, 4 received 7 or 14 days of human-recombinant HGF starting at day 11. Graft function was assessed by daily serum creatinine and biopsies. Immunologic assays, including CD4/CD25 DP and FoxP3+ cells and development of antidonor antibodies, were performed. RESULTS: Without HGF, all six CLAWN recipients developed severe acute rejection (Cre >9 mg/dL) within 3 weeks of transplantation. In contrast, in the four animals that received HGF for 7 to 14 days, stable renal function was observed for more than 50 days, although all grafts were ultimately rejected by postoperative day 80. Percent FoxP3+ cells in the CD4+CD25+ double positive population (T regulatory cells) in peripheral blood monocyte cells decreased in recipients with FK506 induction monotherapy while no reduction was observed in recipients treated with FK506 and HGF. CONCLUSION: This study demonstrates that in CLAWN swine treated with a dose of FK506 insufficient to induce tolerance across a fully MHC mismatched barrier, a short course of HGF may inhibit acute rejection while maintaining T regulatory cells. To our knowledge, this study provides the first evidence in a large animal transplantation model of HGF's immunoprotective effects.