Inhibition of intimal hyperplasia by photodynamic therapy using photofrin.

Photodynamic therapy using photofrin and light energy inhibits human myofibroblast proliferation in cell culture. The purpose of this study is to evaluate its influence on intimal hyperplasia in vivo. Twenty New Zealand White rabbits underwent a standardized intimal injury to both common carotid arteries with a 2 Fr balloon catheter. One week later, half of the animals received photofrin (5 mg/kg) intravenously. The remaining 10 rabbits received no photofrin. Two days later, all neck incisions were reopened and a 1-cm segment of each of the 40 carotid arteries was exposed for 5 min to 80 mW of 630 nm light energy from a continuous wave tunable dye laser (fluence = 7.6 J/cm2). All vessels were harvested 5 weeks post-laser treatment following in vivo fixation with formalin. From each artery, separate cross-sections taken from both the lasered and non-lasered regions of each vessel were mounted and stained for histologic evaluation. Analyzed segments were then divided into four different treatment groups: group I segments consisted of arterial cross-sections which were taken from vessel regions that were injured but received neither photofrin nor laser treatment (group I, n = 20); group II segments also did not receive photofrin but were exposed to light energy (group II, n = 20); group III segments received photofrin but no light energy (group III, n = 20); and cross-sections in group IV were taken from those segments which received both photofrin and laser treatment. Using planimetry, the ratio of the area of intimal hyperplasia (IH) to the area enclosed by the internal elastic lamina (IEL) was measured for each specimen (IH/IEL).(ABSTRACT TRUNCATED AT 250 WORDS)     

Liposomal encapsulation significantly enchances the immunosuppressive effect of tacrolimus in a discordant islet xenotransplant model

BACKGROUND: Encapsulation of tacrolimus (TAC) in a lipid bilayer to form liposome-encapsulated tacrolimus (LTAC) alters the biodistribution profile, half-life, and efficacy in organ allotransplantation models. LTAC has not been applied to either cell transplantation or xenotransplantation. METHODS: To test the efficacy of LTAC in a discordant islet xenograft model, tilapia (fish) islets were transplanted under the left kidney capsules of streptozotocin-diabetic Balb/c mice. Recipient mice (groups I-VI) were treated with: I, untreated; II, empty liposomes; III, TAC (2 mg/kg/day); IV, TAC (5 mg/kg/day); V, LTAC (2 mg/kg/day); or VI, LTAC (5 mg/kg/day); all treatments were for 35 days or until rejection (i.e., two glucose measurements >200 mg/dl). Graft-bearing kidneys were removed for histology after rejection. RESULTS: Mean graft survival time (mGST) for control groups I and II were 6.7+/-1.4 (n=6) and 7.5+/-1.3 days (n=4), respectively. Daily TAC treatment at 2 mg/kg/d (III) did not prolong graft function (mGST=7.7+/-1.6; n=6) although 5 mg/kg/day (IV) produced minimal prolongation to 12.8+/-4.8 days (n=12). Treatment with LTAC at 2 mg/kg/day (V) significantly prolonged mGST to 26.6+/-4.9 (n=5); however, all recipients rejected during treatment (i.e.,<35 days). LTAC at 5 mg/kg/day (VI) further prolonged mGST to 39.9+/-11.8 days (n=12) with only one mouse rejecting before day 35. Histologically, at the time of functional rejection, grafts were generally either totally or partially effaced by mononuclear cells, eosinophils, and fibrosis. In groups VI, islet grafts removed from two mice that died while they were normoglycemic and from a mouse terminated while it was normoglycemic at day 36 were viable, well-granulated, and free from cellular infiltration. The group VI grafts examined at rejection (i.e., 1-2 weeks after discontinuing LTAC) were generally totally obliterated and were in two instances associated with nodular aggregates of atypical lymphocytes resembling posttransplant lymphoproliferative disorder. CONCLUSIONS: LTAC is the most potent immunosuppressive compound we have tested in our discordant fish-to-mouse islet xenograft model; however, toxicity is an issue at high doses.     

Successful transplantation after long-term preservation of dog hearts.

Nucleoside transport inhibition is a new approach to long-term preservation of donor hearts. To evaluate its effectiveness, the following were tested: 1) the effect of nucleoside transport inhibition on high-energy phosphate content after cardioplegic arrest and during long-term cold storage (group I: cardioplegia, control ]n = 18]; group II: cardioplegia plus nucleoside transport inhibitor [n = 18]); 2) the effect of nucleoside transport inhibition on high-energy phosphates and hemodynamic recovery in a modified blood-perfused Langendorff system (group III: 24-h cold storage followed by reperfusion [n = 6]; group IV: addition of nucleoside transport inhibition to cardioplegia but not during reperfusion [n = 6]; group V: addition of nucleoside transport inhibition during reperfusion [n = 6]; group VI: addition of nucleoside transport inhibition to cardioplegia and during reperfusion [n = 6]); and 3) the effect of nucleoside transport inhibition added to cardioplegia and during reperfusion on high-energy phosphate content and outcome after heart transplantation (group VII: no nucleoside transport inhibitor in cardioplegia and during reperfusion [n = 8]; group VIII: addition of nucleoside transport inhibition to cardioplegia and during reperfusion [n = 8]). The following results were obtained: 1) addition of nucleoside transport inhibition prevented high-energy phosphate depletion during cold storage: after 24 h, adenosine triphosphate content in group I was 9.4 +/- 3.1 mumol/g versus 17.7 +/- 3.6 mumol/g dry weight in group II (P less than 0.05); 2) addition of nucleoside transport inhibition to cardioplegia and during reperfusion resulted in greater high-energy phosphate content (adenosine triphosphate in group III was 7.9 +/- 3.5 mumol/g vs. 17.8 +/- 2.8 mumol/g in group VI [P less than 0.05]) and improved hemodynamics upon reperfusion (hearts in group III did not recover, maximum isometric left ventricular pressure development was 1,635 +/- 577 mmHg/sec in group IV, 1,915 +/- 423 mmHg/sec in group V, and 2,437 +/- 201 mmHg/sec in group VI [P less than 0.05, group VI vs. groups IV and V]); and 3) hearts treated with nucleoside transport inhibition in cardioplegia and during reperfusion (group VIII) could be transplanted successfully in contrast to group VII hearts. These data indicate that nucleoside transport inhibition in dogs is highly effective in long-term preservation of donor hearts.     

Effects of levonorgestrel butanoate alone and in combination with testosterone buciclate on spermatogenesis in the bonnet monkey

The spermatogenic effects of levonorgestrel butanoate were studied in adult male bonnet monkeys when administered alone and in combination with testosterone buciclate. Levonorgestrel butanoate (0.25, 1.0 and 2.5 mg kg(-1)) given as two injections on days 0 and 60 (groups II, III, IV) resulted in thickening and folding of the basement membrane and disruption of cell associations in groups III and IV (on day 120). In group II, no apparent changes in testicular histology were observed. When these doses of levonorgestrel butanoate were combined with 40 mg of testosterone buciclate (groups V, VI, VII), maximum changes were seen in group VI in which all stages of spermatogenesis were absent on day 120 except for a small number of spermatogonia. The changes caused by lower dose (group V) and higher dose (group VII) of levonorgestrel butanoate were less prominent than in group VI. A significant decrease in the number of dark A (Ad) and B spermatogonia was observed in all groups except for Ad spermatogonia on day 120 in group V, B spermatogonia on day 60 in group IV and B spermatogonia on day 120 in group III. A significant decrease in pachytene spermatocytes was seen on day 120 in groups V only. Early spermatids showed a significant decrease only in groups V and VII on day 120 of treatment. Advanced spermatids were suppressed significantly in group IV on day 60 and in groups IV and V on day 120. These data indicate that levonorgestrel butanoate (1.0 mg kg(-1)) in combination with 40 mg of testosterone buciclate was the most effective treatment in suppressing spermatogenesis. The site of action of this combination regimen is at the level of renewing Ad spermatogonia.     

Efficacy of corticosteroids in suppression of intimal hyperplasia

The effect of steroids and immunosuppression on the development of intimal hyperplasia was studied in 24 New Zealand white rabbits. Staged bilateral arteriotomy and stripping of intima of the common carotid artery was performed by means of a 2F balloon catheter under 700 mm Hg of pressure. At 3 months, after the control side artery was harvested (N = 24), the rabbits were assigned to three groups of eight animals each: (1) dexamethasone 0.1 mg/kg, (2) cyclophosphamide (Cytoxan) 1 mg/kg, or (3) azathioprine 1 mg/kg intramuscularly. The animals were treated on the day before the contralateral intimal stripping and then were treated six times a week for 8 weeks; the vessels were harvested at 3 months. Twelve-week patency rates were 62.5% in the control group, 83.3% in the dexamethasone group, 100% in the Cytoxan group, and only 33.3% in the azathioprine group. The ratio of the luminal area to the area enclosed by the internal elastic lamina was used as an index of intimal hyperplasia, with a higher ratio indicating less intimal thickening. This ratio of the patient vessels was 0.74 +/- 0.14 (N = 15) for the controls, 0.79 +/- 0.11 (N = 6) for the Cytoxan group, and 0.91 +/- 0.06 (N = 5, p less than 0.05) for the dexamethasone group, which is statistically significant by means of analysis of variance. Occluded vessels had evidence of organized thrombus without any intimal hyperplasia. The decrease in intimal hyperplasia seen in the steroid group suggests a potential role for steroids in small vessel revascularization, but further studies are required.     

Allopurinol/uricase and ibuprofen enhance engraftment of cardiomyocyte-enriched human embryonic stem cells and improve cardiac function following myocardial injury.

OBJECTIVE: A major limitation of stem cell transfer is early donor-cell death. Here, we seek to enhance myocardial repair following injury through transplantation of cardiomyocyte-enriched human embryonic stem cells (hESC) and recipient treatment with cytoprotective (allopurinol+uricase) and anti-inflammatory (ibuprofen) agents. METHODS: We injected 10(6) (15% hESC-derived cardiomyocytes) green fluorescent protein (GFP+) hESC in the infarcted area following left anterior descending artery (LAD)-ligation in SCID-beige mice. In Group I, 1.6 mg allopurinol and 0.2 mg of uricase were injected i.p. for 3 days prior to cell transplantation. In Group II, 0.35 mg/ml of ibuprofen were added to the drinking water before and after cell implantation. In Group III, the LAD was ligated and allopurinol/uricase was administered without cell treatment. In Group IV, ibuprofen was added to the drinking water and the LAD was ligated without additional cell treatment. In Group V, only cells were transplanted. Group VI involved infarcted controls and Group VII involved sham-operated mice (all groups: n=5). We evaluated heart function (ejection fraction (EF)) by MRI (4.7 T) 3 weeks later. The hearts were harvested for histology. RESULTS: Differentiated hESC formed clusters and expressed alpha-sarcomeric actin and Connexin 43. Cell treatment improved heart function, which was best in the ibuprofen- and allopurinol-treated groups (+cell transfer), compared to the infarcted controls [EF: Group I: 76.6+/-8.6%, Group II: 78.6+/-7.3%, Group III: 58.1+/-5.7%, Group IV: 53.9+/-5.2%, Group V: 57.7+/-7.5%, Group VI: 43.5+/-4.3%, and Group VII: 66.3+/-7.8%]. We did not observe tumors in any group. CONCLUSIONS: Allopurinol/uricase and ibuprofen enhance differentiated hESC-engraftment and myocardial restoration following transplantation into the injured heart.     

Suppression of intimal hyperplasia in a rabbit model of arterial balloon injury by enalaprilat but not dimethyl sulfoxide

Intimal hyperplasia appears to result from the deposition of collagen and matrix by medial myofibroblasts, which are stimulated in response to vascular injury. We hypothesized that pharmacologic inhibitors of fibroblast proliferation would suppress the development of intimal hyperplasia. We evaluated the effect of two agents known to inhibit fibroblast proliferation in vitro: enalaprilat, an angiotensin-converting enzyme (ACE) inhibitor, and dimethyl sulfoxide (DMSO), an organic solvent. Thirty-five New Zealand white rabbits underwent standardized balloon catheter injury of the left common carotid artery. Experimental groups received daily intramuscular injections of the following: group I (n = 15), saline solution; group II (n = 10), 0.07 mg/kg enalaprilat; and group III (n = 10), 2 ml/kg of a 25% by weight DMSO solution. Injections were started 1 day prior to injury and continued 5 days a week for 8 weeks. Carotid arteries were perfusion-fixed at 12 weeks and cross-sectioned for measurement by planimetry. Intimal hyperplasia was measured as the ratio of the absolute area of intimal hyperplasia to the normalized area enclosed by the internal elastic lamina (IH/IEL) and was expressed as a percent. Mean values for IH/IEL were as follows: group I (control), 20.6 ± 2.3%; group II (enalaprilat), 9.5 ± 0.7%; and group III (DMSO), 17.6 ± 2.6%. Enalaprilat-treated animals demonstrated a statistically significant suppression of intimal hyperplasia compared with controls (p<0.01, ANOVA, Student's t test), whereas the DMSO-treated group did not. We conclude that enalaprilat is effective in suppressing the development of intimal hyperplasia in this model of arterial injury. These results support the theory that the smooth muscle cell mitogen angiotensin II plays an important role in the proliferation of medial myofibroblasts following vascular injury.Supported in part by a grant from the Joash Foundation and the Veterans Administration.Presented at the Thirty-fifth Annual University Surgical Residents' Conference, Society of University Surgeons, Seattle, Wash., February 13, 1993.We would like to acknowledge the support of Merck Sharp & Dohme of West Point, Pa., for generously supplying the enalaprilat (Vasotec I.V.) used in this study.     

Folate supplementation inhibits intimal hyperplasia induced by a high-homocysteine diet in a rat carotid endarterectomy model

OBJECTIVE: Hyperhomocysteinemia has been implicated as a causative factor in intimal hyperplasia development. The addition of dietary folate in a hyperhomocysteinemia, carotid endarterectomy rat model is postulated to decrease plasma homocysteine levels and, in turn, reduce post-carotid endarterectomy intimal hyperplasia. METHODS: Each rat was fed one of six diets: (1) lab chow with no folate (n = 7), (2) lab chow with 10 mg/kg folate added (n = 3), (3) lab chow with 25 mg/kg folate added (n = 3), (4) a homocysteine diet with no folate (n = 7), (5) a homocysteine diet with 10 mg/kg folate added (n = 5), or (6) homocysteine diet with 25 mg/kg folate added (n = 5). Each rat then underwent an open carotid endarterectomy. In 2 weeks, intimal hyperplasia in the carotid artery was measured. Plasma homocysteine and folate levels were measured. RESULTS: Plasma folate levels rose with folate administration. Plasma homocysteine in the lab chow group was 5.4 +/- 0.5 micromol/L and did not change with the addition of folate. In the homocysteine diet group, plasma homocysteine rose 10-fold over the lab chow group (51.9 +/- 6.5 vs 5.4 +/- 0.5, micromol/L, P <.0001). In the group fed a homocysteine diet with 10 mg/kg folate added, a significant decrease in plasma homocysteine was observed (17.5 +/- 8.5 vs 51.9 +/- 6.5, micromol/L, P =.0003). In the group fed a homocysteine diet with 25 mg/kg folate added, plasma homocysteine levels were further reduced to levels seen in the lab chow group (12.6 +/- 2.6 vs 5.4 +/- 0.5, micromol/L, P = not significant). The relationship between plasma folate and homocysteine was inverse (R = 0.39, P =.0036). Luminal stenosis due to intimal hyperplasia was minimal in lab chow groups and unaffected by folate. The homocysteine diet group demonstrated post-carotid endarterectomy luminal stenosis due to intimal hyperplasia (60.9% +/- 9.2%). In the group fed a homocysteine diet with 10 mg/kg folate added, intimal hyperplasia was reduced, compared with the homocysteine diet group (32.6% +/- 7.4% vs 60.9% +/- 9.2%, P =.009). In the group fed a homocysteine diet with 25 mg/kg folate added, intimal hyperplasia was reduced to lab chow group levels (10.8% +/- 0.8% vs 4.8% +/- 1.0%, P = not significant) and was reduced, compared with the group fed a homocysteine diet with 10 mg/kg folate added. CONCLUSION: The use of folate in this hyperhomocysteinemia carotid endarterectomy model and the resultant attenuation of plasma homocysteine elevation and intimal hyperplasia development lend strong support to homocysteine being an independent etiologic factor in post-carotid endarterectomy intimal hyperplasia.     

Effects of Immunosuppressive Drugs on Rat Renal Ischemia Reperfusion Injury

Introduction

Recent evidence has demonstrated that the immune response and, more specifically, lymphocytes (T and B) and dendritic cells participate as mediators of renal ischemia reperfusion injury (IRI). The aim of this study was, therefore, to evaluate the effect of various immunosuppressive drugs with known activity to prevent IRI among rats undergoing a scheme that is potentially applicable in the clinic.

Methods

Male Sprague-Dawley rats (200-300 g) underwent 60 minutes of ischemia by renal artery clamping and contralateral nephrectomy. The experimental groups (n = 6-7) were as follows: I, Sham; II, Control; III, Rapamycin (R; 1 mg/kg); IV, Methylprednisolone (M; 15 mg/kg); V, Vitamin D3 (VD3; 2 μg/kg); VI, VD3 (1 μg/kg); and VII, M (15 mg/kg) + R (1 mg/kg). Each drug was administered in 2 doses at 6 hours and 1 hour before surgery. Creatinine (Cr) was determined on days 0.1, 2, 3, 5, and 7, and Cr clearance was determined on days 3 and 7. At 7 days nephrectomy was performed to obtain samples for histology to evaluate the degree of acute tubular necrosis.

Results

Mortality from renal insufficiency was between 0 and 33%, except in group V (66%; 4/6; P = .01). Kidney function was similar to controls in all groups except for creatinine at 7 days between group VI (VD3) and control (1.05 vs 0.65; P < .05) but no difference in Cr clearance. Histologically moderate to severe renal damage was greater in groups V and VI (VD3) than controls (P = .04).

Conclusion

We observed that none of the drugs conferred protection against IRI in a time setting relevant to kidney transplantation. Controversy exists regarding R, because some prior studies have shown a deleterious effect on IRI injury, although we did not observe any deleterious effect.     

Prevention of pain on injection of propofol: a comparison of remifentanil with alfentanil in children

AIM: Propofol has a high incidence of pain on injection, particularly when a vein on the back of hand is used. Administration of lidocaine, either before or mixed with propofol remains the most widely used method to attenuate this pain. The use of opioids such as alfentanil and fentanyl has been found to decrease pain induced by propofol injection. The purpose of this study was to evaluate the effects of different doses of remifentanil and alfentanil in minimizing the pain caused by propofol. METHODS: In this randomized, double-blind, placebo-controlled study, healthy premedicated children between the age group of 5-12 years admitted for adenotonsillectomy were randomly allocated to one of 6 treatment groups. Group I: remifentanil 0.25 microg kg(-1); Group II: remifentanil 0.50 microg kg(-1); Group III: alfentanil 15 microg kg(-1); Group IV: alfentanil 20 microg kg(-1) 60 s prior to propofol mixed with 1 mL of 0.9% normal saline; Group V: lidocaine 1 mL of 1% (10 mg) added to 100 mg of propofol and Group VI: normal saline. During the injection of propofol (3 mg kg(-1)) pain perception was assessed with a four-point behavioural scale: none, mild, moderate, or severe. RESULTS: There were 52 subjects in Group I, 51 in Group II, 49 in Group III, 52 in Group IV, 52 in Group V and 52 in Group VI; 63.46% of patients in Group I, 39.21% in Group II, 38.77% in Group III, 36.53% in Group IV, 38.46% in Group V and 84.61% in Group VI experienced pain. Statistically, Groups II, III, IV and V were significantly better than placebo in the reduction of propofol pain (P<0.0001). Groups II, III and IV significantly reduced the pain in comparison with Group I (P<0.001). CONCLUSION: Pretreatment with intravenous remifentanil 0.5 microg kg(-1), alfentanil 15 microg kg(-1) and 20 microg kg(-1) were equally effective in reducing pain associated with propofol injection in children between the age group of 5-12 years.     

Dosage and timing of Photofrin for photodynamic therapy of intimal hyperplasia

Photodynamic therapy has been recommended as a method of preventing intimal hyperplasia. The purpose of this study was to determine the dose and timing of Photoftin porfimer sodium needed to achieve a 3:1 or higher ratio between injured and control arteries after balloon endothelial injury. New Zealand White rabbits were anesthetized and their right femoral artery surgically exposed. A 4Fr Fogarty balloon catheter was passed retrograde into the lower abdominal aorta, inflated and pulled distally into the external iliac artery six times. All rabbits received heparin 100 IU/kg. Arteriotomies were closed and the animals recovered. Rabbits (n = 5 per group) were given intravenous Photofrin at a dose and time according to the following scheme: group I, 5.0mg/kg immediately after balloon injury; group II, 2.5mg/kg immediately after injury; group III, 5.0 mg/kg after 1 week; group IV, 5.0 mg/kg after 2 weeks; or group V, 2.5 mg/kg after 2 weeks. Animals were killed 24 h after drug administration and the aortoiliac segments removed for spectrophotofluorometric determination of Photofrin levels from injured and control segments. Mean(s.d.) ratios of injured: control arteries for groups I to V were 4.8 (2.6), 2.8 (1.2), 3.0 (1.0), 1.4 (0.3) and 1.0 (0.0) respectively. This ratio was significantly higher for group I rabbits compared with groups IV and V (P< 0.01, ANOVA). Fluorescence and light microscopy showed that Photofrin was localized primarily in the tunica media, and that the drug must be administered before significant intimal hyperplasia occurs. This study suggested that the Photofrin dose of 5 mg/kg given immediately or within 7-days of injury achieved at least a 3:1 ratio between injured to control arterial segments. Ratios based on this dose were significantly higher than those from rabbits who received Photofrin 2 weeks after endothelial injury.     

Postoperative immunotherapy of murine C1300-neuroblastoma.

Low-dose cyclophosphamide (CY) is an immunomodulating agent that down-regulates T suppressor cell function. This study investigates postoperative immunotherapy with CY as an alternate treatment for advanced immunogenic tumors such as neuroblastoma that typically respond poorly to traditional high-dose chemotherapy. A/J mice with 1.5-cm subcutaneous C1300-neuroblastoma (C1300-NB) tumors were divided into the following treatment groups: I, untreated (n = 14); II, 85% tumor resection (n = 18); III, sham-operated (n = 18); IV, multiple-dose CY (n = 6); V, 85% resection and single-dose CY (n = 14); VI, 85% resection and multiple-dose CY (n = 14). CY (100 mg/kg, intraperitoneally) was given initially 24 hours post-operatively to groups IV, V, and VI. Groups IV and VI also received weekly maintenance doses of 25 mg/kg CY. Results showed significantly increased survival (log-rank test) in CY-treated groups (IV, V, VI) compared with control groups (I,II,III). Cures were observed only in groups receiving partial resection plus CY (V, 7%; VI, 29%). Although surgical debulking of tumor alone (II) did not enhance survival, the procedure normalized depressed total lymphocyte counts and the subpopulation of Lyt 2,3+ (T suppressor/cytolytic cells) in the immediate postoperative period during which immunotherapy with CY was instigated. This may have contributed to the success of CY immunotherapy. To characterize the tumor-host immune interaction, additional studies were performed. Results showed the following. (1) Mice cured by debulking plus CY (from groups V and VI) could not be successfully reimplanted with C1300-NB, demonstrating immunologic mediation by CY.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of aspirin, dipyridamole, and cod liver oil on accelerated myointimal proliferation in canine veno-arterial allografts.

The effects of the administration of aspirin (ASA), dipyridamole (DPM), and cod liver oil (CLO) on graft patency rate and degree of intimal hyperplasia were investigated in a canine, hypercholesterolemic veno-arterial allograft model in an attempt to modify this immunologically mediated vascular injury. The drug regimens were ASA 1 mg/kg/day, DPM 10 mg/kg/day, combined ASA and DPM (ASA + DPM), and CLO (1.8 g/day eicosapentanoic acid [EPA] and 1.2 g/day docosahexanoic acid [DHA]), and control. The early angiographic patency rate (1-3 weeks) was 81% +/- 10% (+/- 70% confidence limits); the 90-day overall patency rate was 60% +/- 4% (87/144), with no statistically significant differences among the groups (range 46 +/- 10-71 +/- 9%). Qualitatively, there was no difference in luminal thrombus, intimal hemorrhage, or lesion eccentricity. Considering the relatively short time of graft implantation, an extensive amount of microscopic disease was observed; quantitatively, the mean intimal thickness was 515 +/- 17 microgram overall but was not statistically different between the groups. The fraction of potential lumenal area occupied by intimal thickening was 0.37 +/- 0.01 but again did not differ significantly between the groups. These doses of ASA, DPM, ASA + DPM, and CLO did not alter graft occlusion or retard the marked degree of subintimal myointimal cell hyperplasia that was generated in this hypercholesterolemic canine veno-arterial allograft preparation. Possible explanations for these negative findings include inadequate dosage or form of omega-3 fatty acids and the antiplatelet drugs administered, excessive variability in graft response due to uncharacterized immunologic histocompatibility, and the possible influence of non-platelet-mediated mechanisms. Nevertheless, this preparation is attractive as a reproducible model of accelerated (immunologically mediated) experimental arteriosclerosis.     

Safe injection of cultured schwann cells into peripheral nerve allografts

The effects of cultured host Schwann cells on axonal regeneration in peripheral nerve allografts were studied. Fischer rats served as recipient animals and Buffalo rats provided nerve allografts. Animals were randomized into 9 groups. Rats receiving tibial nerve isografts were left untreated (group I), or injected with isogeneic Fischer Schwann cells (group II) or placebo suspension (group III). Allografts obtained from Buffalo rats were left untreated (group IV), or received isogeneic Fischer Schwann cells (group V), 2 mg/kg Cyclosporin A and Fischer Schwann cells (group VI), 5 mg/kg Cyclosporin A (group VII), or 5 mg/kg Cyclosporin A with Schwann cells (group VIII). No Schwann cell tumors were identified 4 or 8 weeks postoperatively. Group IX animals, harvested 3 days postoperatively, demonstrated no evidence of injection injury. Schwann cells modestly improved axonal regeneration in both isografts and allografts and may have a clinical role in the treatment of peripheral nerve allografts.     

Mechanical bowel preparation with different solutions in rats with selective left colonic ischemia and reperfusion injury

BACKGROUND: The aim of this study was to evaluate the effects of preoperative mechanical bowel preparation (MBP) on colonic ischemia/reperfusion (I/R) injury. METHODS: Seventy adult male Sprague-Dawley rats were divided randomly into 7 equal groups of 10 rats each. Groups were assigned as follows: group I = sham surgery; group II = I/R of left colon (control group); group III = intravenous heparin and metronidazole followed by I/R of the left colon; groups IV through VII = before I/R of the left colon, heparin and metronidazole and MBP were performed with sodium chloride (NaCl), Na phosphate, polyethylene glycol, and mannitol, respectively. Histopathologic and biochemical parameters were evaluated. RESULTS: According to the histopathologic changes, the groups least affected by I/R injury were groups V and VII. Catalase activity was significantly higher in groups V and VII, and copper-zinc superoxide dismutase activity was significantly higher in group VII compared with the control group (P <.002). CONCLUSIONS: MBP with sodium phosphate and mannitol appears to be more protective against I/R injury.     

Propofol prevents delayed neuronal death following transient forebrain ischemia in gerbils

PURPOSE: This study was conducted to ascertain whether propofol may protect against delayed neuronal death in the hippocampal CA1 subfield in gerbils. METHODS: Thirty-five gerbils were randomly assigned to five groups: Group I, the control group, a sham operation treated with physiological saline solution (PSS); Group II, ischemia/reperfusion treated with PSS; Group III, ischemia/reperfusion treated with 50 mg x kg(-1) propofol; Group IV, ischemia/reperfusion treated with 100 mg x kg(-1) propofol; Group V ischemia/reperfusion treated with 150 mg x kg(-1) propofol. Transient forebrain ischemia was induced by occluding the bilateral common carotid arteries for four minutes under N2O/O2/halothane anesthesia after administration of propofol or PSS. Five days later, histopathological changes in the hippocampal CA1 subfield were examined using a light microscope and degenerative ratio of the pyramidal cells were measured according to the following formula: (number of degenerative pyramidal cell/total number of pyramidal cells per 1 mm of hippocampal CA1 subfield) x 100. RESULTS: In group II, the pyramidal cells were atrophic and pycnotic; vacuolation and structural disruption of the radial striated zone was observed. In the other four groups, these changes were not observed. The degenerative ratios of pyramidal cells were as follows; group I: 5.9 +/- 1.9%, group II: 94.6 +/- 2.5% (P < 0.01), group III: 10.7 +/- 1.7%, group IV: 9.7 +/- 1.8%, group V: 9.2 +/- 1.9%. CONCLUSION: This study suggests that propofol may prevent delayed neuronal death in the hippocampal CA1 subfield after cerebral ischemia/reperfusion in gerbils.     

Evaluation of the ability of levonorgestrel butanoate alone, or in combination with testosterone buciclate, to suppress spermatogenesis in the bonnet monkey (Macaca radiata)

Levonorgestrel butanoate, 0.25, 1.0 and 2.5 mg/kg, administered as two injections 60 days apart (groups II, III, IV), failed to suppress spermatogenesis consistently and uniformly in adult bonnet monkeys (group size, n=6) compared to controls (group I). Levonorgestrel butanoate at the same doses combined with two simultaneous injections of 40 mg testosterone buciclate (groups V, VI, VII), consistently suppressed spermatogenesis in the period 60-240 days and in most animals to azoospermia or severe oligozoospermia (<5 x 106/mL) during days 90-210. The degree and duration of suppression were greatest in group VI. Sperm motility declined in all treated animals and spermatozoa in the semen of animals from groups V and VI lost all progressive motility in the period 60-150 and 60-210 days, respectively. The changes in testosterone levels were similar in groups V and VI, increasing within 24 h after the combined injection to reach a peak by day 28 followed by a sharp decrease until day 67. The second injection increased testosterone levels by a lesser degree to peak levels on day 81. In group VII, testosterone levels decreased until day 59 after the first injection but increased to a maximum on day 81 after the second injection followed by a gradual decrease until day 150 to below baseline values. Peak levels of serum levonorgestrel were observed 1-7 days after injection of levonorgestrel butanoate alone. Clearance of the drug was slow, being detectable in the circulation until day 330 of the 360 day study period in the high dose group. Dose-response increases to peak levels of levonorgestrel were attained on day 7 in groups V, VI and VII, after the first injection. After the second injection, peak levels were seen on day 61 in groups V and VI and on day 81 in group VII. Levonorgestrel was no longer detectable in blood in groups V and VI by days 210 and 300, respectively, but small circulating amounts remained in group VII at the conclusion of the study on day 360. This study indicates that when levonorgestrel butanoate is combined with a long-acting androgen and injected at two-monthly intervals, effective and reversible suppression of spermatogenesis is achieved.     

Polyphenol (-)-epigallocatechin gallate protection from ischemia/reperfusion-induced renal injury in normotensive and hypertensive rats

INTRODUCTION: The effect of epigallocatechin gallate (EGCG) in an in vivo renal model of ischemia with reperfusion (I/R) was compared between normotensive (WKR) and hypertensive (SHR) rats. METHODS: WKR (groups I, II, III) and SHR groups (groups IV, V, VI) were divided into three types. Groups I and IV were sham-operated animals; groups II and V were subjected to 45 minutes of renal I/R; and groups III and VI received 10 mg/kg EGCG intravenously at the time of reperfusion. Three days after renal I/R, we compared renal function markers, malondialdehyde (MDA), and histologic changes. RESULTS: Following renal I/R, levels of blood urea nitrogen (BUN) and serum creatinine (sCr) were increased and serum creatinine clearance (CrCl) decreased in group V compared to group II (P < .001). Those receiving EGCG treatment (groups III and VI) had decreased BUN and sCr compared to non-EGCG I/R groups (P < .001), but not surprisingly, higher than sham groups. CrCl was lowest in the SHR groups. The MDA was significantly decreased after EGCG treatment (P = .028 in group III, P = .002 in group VI). Following renal I/R, tissue necrosis was more severe among SHR (P < .001). However, the ratio of regeneration to damage significantly increased in SHR after EGCG treatment. CONCLUSIONS: The reperfusion injury was greater among SHR compared with WKR in terms of renal function, lipid peroxidation, and tissue damage. EGCG treatment significantly ameliorated renal impairment and promoted tissue regeneration following renal I/R.     

Inhibition of myointimal hyperplasia and macrophage infiltration by estradiol in aorta allografts.

A major cause of organ graft loss after heart transplantation is accelerated atherosclerosis. In this study we used aorta allografts and investigated the effect of estradiol-17 beta treatment on both the degree of myointimal hyperplasia and morphological changes evaluated by light and electron microscopy. Outbred New Zealand white male rabbits (2.7-3.5 kg) were fed cholesterol (0.5%) from one week prior to transplantation, and until sacrifice three weeks later. The donor abdominal aorta was transplanted end-to-end to the right carotid artery of the recipient animals. Immediately following surgery, cyclosporine (10 mg/kg/d s.c.) was administered to prevent graft rejection. The allograft recipients were randomly assigned to one of five groups and treated with cottonseed oil (placebo) or estradiol cypionate at 1, 10, 100, or 1000 micrograms/kg/d i.m. for 3 weeks. The aorta grafts were harvested and fixed for transmission electron microscopy and morphometry. The area of myointimal thickening was calculated as a percent of total vessel area (mean +/- SEM); the control group was 6.6 +/- 0.5% (n = 5). Estradiol treatment significantly inhibited (P less than 0.05) myointimal hyperplasia at all doses. The values were 3.9 +/- 0.6% (n = 6) for 1 microgram/kg/day; 4.4 +/- 0.7% (n = 5) for 10 micrograms/kg/day; 3.5 +/- 0.4% (n = 6) for 100 micrograms/kg/day; and 2.9 +/- 0.1% (n = 3) for 1000 micrograms/kg/day. Electron microscopic evaluation revealed that the four doses of estradiol protected the endothelium from the degenerative changes seen in all aorta allografts from the animals in the control group. Furthermore 10, 100, and 1000 micrograms/kg/day of estradiol prevented the appearance of vacuolized macrophages (foam cells) and also the vacuolization of smooth muscle cells that was observed in the aorta allografts from the control group and the group treated with 1 microgram/kg/day of estradiol. We conclude that the inhibitory effect of estradiol on the development of graft atherosclerosis may be due to inhibition of smooth muscle cell proliferation and preservation of ultrastructurally normal endothelial cells. The inhibitory effect on foam cell production and a concomitant vacuolization of smooth muscle cells may play a lesser role. We suggest that estrogen replacement therapy may be beneficial in postmenopausal women with organ allografts.     

The role of aspirin and dipyridamole on vascular DNA synthesis and intimal hyperplasia following deendothelialization

Platelets are implicated both in acute thrombotic events and, through platelet-derived growth factor, in the development of intimal hyperplasia. We have investigated, in vivo, the influence of aspirin and dipyridamole on vascular smooth muscle cell proliferation and DNA synthesis following balloon catheter injury. Fifty-eight male, New Zealand white rabbits were divided equally into two groups; the test group was fed aspirin (14 mg/kg/day) and dipyridamole (9 mg/kg/day) from 2 days prior to surgery until sacrifice at 1, 2, 3, 4, 7, 14, or 28 days after injury. All animals were sacrificed 1 h after injection of [3H]thymidine and the smooth muscle cell DNA specific activity and total kinetic activity were determined. Intimal hyperplasia was measured by light microscopy and intimal nuclear proliferation was determined by counting nuclei per millimeter of internal elastic lamina. Nuclear proliferation was maximal at 14 days (25 +/- 1.2) but intimal hyperplasia was still increasing at 28 days. DNA specific activity after 24 hr (test: 4 +/- 2 dpm/micrograms DNA; control: 3.3 +/- 3 dpm/micrograms DNA) was similar to basal levels in uninjured rabbits. DNA synthesis peaked in both groups between the second and third day (test: 177 +/- 27 dpm/micrograms DNA; control: 185 +/- 39 dpm/micrograms DNA) and then declined slowly toward baseline values. There was no significant difference between treated and normal rabbits in either [3H]thymidine incorporation, nuclear proliferation, or development of intimal hyperplasia despite 90% inhibition of platelet aggregation and a significant reduction (78%) in [14C]serotonin release following collagen challenge (6 micrograms/ml).(ABSTRACT TRUNCATED AT 250 WORDS)