Effect of tolmetin sodium dihydrate on adhesion formation by intraperitoneal administration of antineoplastic agents.

Antineoplastic agents are currently being administered through catheters placed intraperitoneally to treat cancer localized to the peritoneum. This route allows for high local concentrations of antineoplastic drug at the tumor site with low levels of the drug systemically, thereby reducing the systemic toxicity. However, there are complications with this mode of delivery, including a decrease in catheter patency and induction of adhesion formation, which leads to decreased drug dispersion and limits continuing drug administration. A model was developed in rats to mimic this method of antineoplastic drug administration that produced fibrin deposition around the catheter and adhesion formation involving bowel, intestines and liver. All antineoplastic agents tested, including Adriamycin, methotrexate, bleomycin, mitoxantrone and cisplatin, induced moderate to severe adhesion formation with varying effects on catheter patency. When an intraperitoneal bolus of tometin encapsulated in liposomes was tested with Adriamycin delivered via an osmotic minipump, a reduction in adhesion formation was observed. However, highly significant adhesion reduction was found when tolmetin was coadministered with the antitumor agents.

Thyroid hormone-induced GLUT-1 expression in rat cerebral tissue: effect of age.

To determine if thyroid hormone (TH) modulates the expression of cerebral glucose transporter one (GLUT-1) and whether there are age-related differences in TH effect, young male Fischer 344 rats (6 months old) and aged rats (26 months old) were studied at euthyroid, hyperthyroid and hypothyroid conditions. Immunoblot analysis indicated that 55 kDa GLUT-1 mass was decreased in hypothyroid young rats (174 +/- 15 arbitrary units) and hyperthyroid rats (144 +/- 22) compared to euthyroid young rats (395 +/- 57) P < 0.01. Jr. aged rats the 55 kDa GLUT-1 mass was significantly increased in hyperthyroidism (392 +/- 49) compared to euthyroid aged rats (237 +/- 27) P < 0.05. The 45 kDa isoform of GLUT-1 in rat cerebral tissue did not significantly change with age or thyroidal state. The changes in 55 kDa GLUT-1 mass did not correlate with the changes of GLUT-1 mRNA content. It is concluded that alterations in cerebral GLUT-1 content in response to altered thyroid state are age-specific.

Immunocytochemical localization of the low-affinity nerve growth factor receptor (p75NGFR) in the cuneate nucleus of the rat and its relationship to cytochrome-oxidase activity.

Immunohistochemical staining for the 75-kDa, low-affinity nerve growth factor receptor (p75NGFR), within the cuneate nucleus (CN) of the adult rat revealed that this receptor is concentrated rostrocaudally in the middle CN (approximately 0.2-0.9 mm caudal to the obex), corresponding to that portion of the CN receiving densest projections of cutaneous primary afferent terminals. Furthermore, dense patches of p75NGFR-like immunoreactivity appear to correspond to the 'blotches' of cytochrome-oxidase activity observed in the middle region of the CN. This close correspondence between the localization pattern of p75NGFR in the CN and its functional organization suggests an important role for trophic factors in the CN's development and/or maintenance.

Reduction of adhesion formation by intraperitoneal administration of Arg-Gly-Asp-containing peptides.

OBJECTIVE: To evaluate the ability of a variety of peptides containing the Arg-Gly-Asp (RGD) sequence to reduce the formation of intraperitoneal adhesions in a rabbit model. DESIGN: Prospective, randomized, double-blinded study. SETTING: University-based laboratory. ANIMAL(S): New Zealand white rabbits. INTERVENTION(S): Administration of RGD-containing peptides. MAIN OUTCOME MEASURE(S): Reduction of adhesion information. RESULT(S): In initial studies, two RGD-containing peptides (3 or a 10 amino acid peptides) were administered via Alzet miniosmotic pump to the site of injury. Administration of either of these peptides significantly reduced adhesion formation, but the larger peptide was more efficacious and reduced variability in the response. Further studies then were conducted with RGD-containing peptides five to six amino acids in length. Administration of these peptides also significantly reduced adhesion formation in a standard rabbit model. Administration of three of these peptides in a viscous vehicle at the end of surgery was also effective in reducing adhesion formation. CONCLUSION(S): The most effective combination tested was RGD-containing peptide Gly-dser-Arg-Gly-Asp-Ser-Pro in a viscous, cremophor-containing vehicle. These studies demonstrate that administration of an RGD-containing peptide was effective in reducing adhesion formation in this model.

Reduction of adhesion formation with hyaluronic acid after peritoneal surgery in rabbits.

OBJECTIVE: To examine the effect of hyaluronic acid, a high-molecular-weight glucosaminoglycan found in the extracellular matrix, on the formation of adhesions, a major source of postoperative complications. DESIGN: The ability of hyaluronic acid to reduce adhesion formation was evaluated using a standardized rabbit model. The material was administered i.p. at the end of surgery. SETTING: University laboratory. ANIMAL(S): New Zealand White female rabbits. INTERVENTION(S): Intraperitoneal administration of various formulations of hyaluronic acid at the end of surgery. MAIN OUTCOME MEASURE(S): One week after surgery, a second laparotomy was performed and the extent of adhesion formation was determined. RESULT(S): Five separate molecular weight ranges of hyaluronic acid representing eight viscosities between 1,000 and 12,000 centipoise (CPS) were shown to reduce adhesion formation in this model. All volumes, 1 to 30 mL, of hyaluronic acid tested reduced adhesion formation. In addition, the low-viscosity, low-molecular-weight hyaluronic acid significantly reduced adhesion formation when added to the trauma site or when injected at a site remote from the trauma area. CONCLUSION(S): This study showed that hyaluronic acid administered at the end of surgery reduced adhesion formation.

Reduction of adhesion formation by intraperitoneal administration of anti-inflammatory peptide 2.

Adhesion formation is a major source of postoperative morbidity and mortality. Therefore, the reduction of postoperative adhesion formation would be of clinical benefit. Various modalities have been shown to reduce adhesion formation, including fibrinolytic enzymes, nonsteroidal anti-inflammatory drugs, and barriers that reduce the apposition of sites of potential adhesion formation. This study examined the ability of a phospholipase A2 inhibitor, anti-inflammatory peptide 2 (antinflammin), to reduce the formation of intraperitoneal adhesions in two rabbit models of adhesion formation. In the sidewall model, antinflammin was administered via Alzet miniosmotic pump for the entire postoperative interval, and there was a dose-dependent reduction in the area of the sidewall injury that was involved in adhesions to the cecum and the bowel. In the double uterine horn model, antinflammin was administered via Alzet miniosmotic pump to the area of injury for either 1, 2, 3, or 7 days. Administration of antinflammin for as little as 24 h after surgery significantly reduced the extent of adhesion formation. Administration of the peptide for longer periods of time did not further increase the reduction in adhesion formation. These studies clearly demonstrate that postoperative administration of antinflammin to the site of injury reduced the formation of postoperative adhesions in two animal models.

Reduction of adhesion formation by intraperitoneal administration of a recombinant Hirudin analog.

Adhesion formation is a major source of postoperative morbidity and mortality. Therefore, the reduction of postoperative adhesion formation would be of clinical benefit. Various modalities have been shown to reduce adhesion formation, including fibrinolytic enzymes, nonsteroidal anti-inflammatory drugs, and barriers that reduce the apposition of sites of potential adhesion formation. This study examined the ability of an inhibitor of thrombin, a recombinant hirudin analog (recHirudin), to reduce the formation of intraperitoneal adhesions in two rabbit models of adhesion formation. In the sidewall and double uterine horn models, recHirudin was administered via Alzet miniosmotic pump for the entire postoperative interval. In both of these models, there was a dose-dependent reduction in adhesion formation as measured by (1) the area of the sidewall injury that was involved in adhesions to the cecum and the bowel or (2) the involvement of the uterine horns to themselves or other peritoneal organs. These studies clearly demonstrate that postoperative administration of recHirudin to the site of injury reduced the formation of postoperative adhesions in two animal models.

Prevention of adhesion formation with intraperitoneal administration of tolmetin and hyaluronic acid.

Adhesion formation after peritoneal surgery is a major source of postoperative complications and pain. Previous studies showed that intraperitoneal administration of the nonsteroidal anti-inflammatory drug tolmetin reduced adhesion formation after two types of peritoneal surgery. The effect of tolmetin combined with hyaluronic acid (HA), a high-molecular-weight glucosaminoglycan found in the extracellular matrix, on the formation of adhesions was examined. In this study, the effect of tolmetin in HA on adhesion formation was evaluated in a standardized rabbit model. The medicament was administered intraperitoneally at the end of surgery. One week after surgery, a second laparotomy was performed and the extent of adhesion formation was determined. A range of molecular weights (7.5 x 10(5)-2 x 10(6) Da) and viscosities (1000-25,000 centapoise) of HA in combination with tolmetin was effective in reducing adhesion formation. However, low viscosity HA solutions in combination with tolmetin, 0.5-2.0 mg/mL, were most efficacious in reducing adhesion formation. These data suggest that HA, in combination with tolmetin, acts as an effective carrier to reduce adhesion formation in the abdominal cavity after surgery.

Reduction of adhesion formation by intraperitoneal administration of various anti-inflammatory agents.

Adhesion formation is a major source of postoperative morbidity and mortality. Therefore, the reduction of postoperative adhesion formation would be of clinical benefit. Various modalities have been shown to reduce adhesion formation, including fibrinolytic enzymes, nonsteroidal anti-inflammatory drugs, and barriers that reduce the apposition of sites of potential adhesion formation. In this report, the ability of three compounds with different mechanisms of action, all-trans-retinoic acid, quinacrine, and dipyridamole, to reduce the formation of intraperitoneal adhesions was examined in two rabbit models. In the sidewall model, the medicaments were administered via an Alzet miniosmotic pump for the entire postoperative interval. With all three agents, there was a reduction in the area of the sidewall injury that was involved in adhesions to the cecum and the bowel at both doses tested. In the same model, quinacrine also reduced the area of the sidewall injury that was involved in adhesions to the cecum and the bowel. At the higher concentrations of quinacrine, there was a deposition and walling off of the quinacrine at the site of delivery. In the double uterine horn model (DUH), the medicaments were administered via an Alzet miniosmotic pump to the area of injury for either 1, 2, 3, or 7 days. Administration of all three compounds for as little as 24 h after surgery significantly reduced the extent of adhesion formation. However, there was a further reduction in the amount of adhesion when the retinoic acid or dipyridamole was administered for 72 h postoperatively. However, when the quinacrine was administered for longer times postoperatively, the amount of adhesion reduction observed was less. These studies demonstrate that postoperative administration of retinoic acid, quinacrine, or dipyridamole to the site of injury reduced the formation of postoperative adhesions in two animal models.

Temporal response of leukocyte accumulation in the thoracic cavity after two types of surgical injury.

To characterize pleural healing, we quantitated leukocyte accumulation in the pleural cavity and histological changes after two types of thoracic surgery. Rabbits underwent intercostal thoracotomy followed by abrasion of the parietal pleura and ligation and resection of the right middle lobe of the lung (group A), or only abrasion of the parietal pleura (group B). After surgery, the influx of leukocytes into the pleural exudate was characterized by an increase in the number of polymorphonuclear neutrophils (PMNs) followed by monocytes/macrophages. In group A, the total number of leukocytes reached maximum levels on days 5-7 after surgery, 80% of which were monocytes/macrophages. In group B, the total number of leukocytes reached peak levels on postsurgical day 3, 85% of which were monocytes/macrophages. Histologically, we observed a relative delay in pleural healing in group A compared with group B. An inflammatory response including appearance of fibrinous exudates and infiltration of acute inflammatory cells occurred in group A on days 1-3 after surgery. On days 5-7, an increase in submesothelial connective tissue was seen. An increase in cellularity was observed in this layer (fibroplasia) and the wound surface was covered by macrophagelike cells. In group B, disappearance of fibrinous exudates and fibroplasia occurred by day 3. In both groups, these histological changes from inflammatory phase to proliferative phase occurred on the day when the number of monocytes/macrophages in the pleural cavity reached peak levels. These data demonstrate that different types of thoracic injury alter the kinetics of leukocyte accumulation in the pleural cavity and the healing process of parietal pleura, suggesting that macrophages that accumulate in the pleural cavity may be implicated in postsurgical repair.

In vivo administration of tolmetin in hyaluronic acid modulates protease levels in postsurgical macrophage-conditioned media.

Tolmetin sodium in a hyaluronic acid carrier (tolmetin-HA) was previously shown to reduce adhesion formation and alter the kinetics and levels of cellular influx into the peritoneal cavity after surgery. In this study, the effect of tolmetin-HA on the level of protease activity in macrophage-conditioned media was determined. The level of collagenase activity in macrophage-conditioned media was suppressed at 12 and 24 h after administration of tolmetin-HA. Alternatively, the peak level of elastase activity measured in macrophage-conditioned media was unchanged after tolmetin-HA treatment, but the kinetics of expression of maximal protease activity was delayed from 12 h in the control surgical rabbits to 24 h in tolmetin-HA-treated rabbits. Elevated plasminogen activator activity was detected in acid-treated conditioned media from the tolmetin-HA-treated rabbits when compared to control levels. However, no alteration in the level of plasminogen activator inhibitor activity was present in conditioned media of macrophages harvested from tolmetin-HA-treated rabbits compared to controls. These data suggest that tolmetin-HA treatment altered the levels of neutral protease activity secreted by postsurgical macrophages and may therefore elevate the fibrinolytic potential of the peritoneal cavity after surgery.

Comparative efficacy of nonsteroidal anti-inflammatory drugs and anti-thromboxane agents in a rabbit adhesion-prevention model.

A variety of nonsteroidal anti-inflammatory drugs (NSAIDs) has been found to inhibit postsurgical peritoneal adhesion formation in a number of animal models. A rabbit uterine horn adhesion model was used to directly compare several commonly used NSAIDs of different chemical classes in a single animal study to evaluate their ability to prevent adhesion formation. The effect of thromboxane inhibitors on adhesion prevention was also evaluated. Each of the NSAIDs tested (tolmetin, ibuprofen, aspirin, and indomethacin) showed significant and comparable efficacy. In this same study, imidazole, a thromboxane synthetase inhibitor, also showed significant efficacy. In a second study, ridogrel, an inhibitor of thromboxane synthetase as well as a thromboxane A2 receptor blocker, also showed significant efficacy in reducing peritoneal adhesion severity. These results further support the view that NSAIDs act to prevent adhesions through a common mechanism. In addition, thromboxane A2 inhibitors were also shown to be efficacious in adhesion prevention, suggesting that platelets may play a substantial role in adhesion formation.

Expression of cyclin protein after thermal skin injury in a guinea pig model.

In this study, an immunohistochemical stain for cyclin was used to quantitate proliferating elements in hair follicles at the edge of and within thermal burn areas. Biopsy specimens from thermal injury in the guinea pig (day 1 through day 28) were sectioned and stained with MIB-1 antibody, which recognizes cyclin, a protein expressed during epithelial cell proliferation. At the edge of the burn, 89 +/- 6.1 (SD) cells per medium power field (x 10, mpf) were MIB-1-positive on days 1 through 16. On day 17, the number of positive cells increased, reaching peak values on days 20 to 28 (271 +/- 12.7 cells/mpf). Within the burn, minimal staining was observed from day 1 to day 15 (12.7 +/- 1.6 cells/mpf). Thereafter the number of MIB-1-positive cells increased and plateaued with an average of 96.4 +/- 9.0 cells/mpf on days 20 through 28. In conclusion, immunohistochemical staining of dermal biopsy specimens with MIB-1 antibody may provide a quantitative method for the evaluation of tissue damage and healing after thermal injury.

Histologic alterations in dermal repair after thermal injury effects of topical angiotensin II.

Previously, we determined that quantitative assessment of epithelialization of the burn site could be performed using quantitative immunohistochemistry with an antibody to the protein cyclin. In this study, the effect of administration of angiotensin II (AII) on two histologic parameters of healing-the number of vascular channels at the burn site and the number of cells proliferating in hair follicles at the edge of the burn and within the burn-were evaluated. Beginning on day 4, vascular channels were noted within the burn site. Significantly more channels were noted in the burns treated with AII than those treated with placebo. With the exception of 3 postinjury days, this increase continued through day 17. Thereafter, the number of vascular channels peaked, and no differences were noted between control and treated burns. The number of cells proliferating in the hair follicle was also evaluated. At the edge of the burn, on average, 126 cells per microscope field (10x) were undergoing proliferation in the AII-treated burn on days 1 through 16 after burn injury. This is approximately a 50% increase over the number of cells proliferating in the placebo-treated burns. On day 12 (approximately 5 days before that observed in control burns), this AII-dependent proliferative response began to increase and peaked on day 19 at a level comparable to control. Thereafter, the proliferative response remained at this level through day 28. Within the area of the burn on days 1 through 15, 21 cells per medium power field on average (approximately 50% more than control) were undergoing proliferation. As on the edge of the burn, an AII-dependent increase in the number of cells proliferating in the hair follicles was observed during the latter phase of healing (on day 16 after the initiation of injury). However, unlike the edge of the burn, administration of AII led to a continued increase (approximately 50%) in the number of cells per field undergoing proliferation. AII increased neovascularization and cellular proliferation after burn injury. Through an increase in these two cellular events, AII may in turn accelerate healing of tissues after thermal injury.

Inhibition of postsurgical adhesions in a standardized rabbit model: intraperitoneal treatment with tissue plasminogen activator.

The direct effects of tissue plasminogen activators (TPA) delivered locally into the peritoneal cavity at the site of injury on the subsequent formation of postsurgical adhesions in rabbits were studied. A 3 x 5 cm flap of parietal peritoneum (approximately 1 mm thick) was sharply dissected from the right lateral peritoneal sidewall. The serosal surface of the adjacent small bowel was abraded with a scalpel until punctate bleeding developed. Alzet minipumps containing varying doses of TPA were sewn into the subcutaneous space with their polyethylene catheter tips leading from the pump into the peritoneal cavity, 2-3 mm over the injury test site. Within 7 days after the implantation of the pumps, the rabbits were killed and the extent of adhesions was determined. The threshold dosage at which significant beneficial antiadhesion properties are obtained was between 0.015 and 0.05 mg TPA/mL of buffer at 6 x 10(-4) mg/kg/day. Effective adhesion prevention occurred with only two days of treatment beginning the day of surgery.

Inhibition of postsurgical adhesions in a standardized rabbit model: II. Intraperitoneal treatment with heparin.

The formation of adhesions after peritoneal trauma is thought to result from the deposition of fibrin and its subsequent organization by fibroblast ingrowth and, in some cases, neovascularization and reepithelialization. Since heparin is an effective anticoagulant, and clotting is a major contributor to postsurgical fibrin deposition, we studied the effects of heparin delivered locally into the peritoneal cavity at the site of the injury on the subsequent formation of postsurgical adhesions. A flap of parietal peritoneum (approximately 1 mm thick) was dissected from the right lateral peritoneal sidewall of New Zealand white female rabbits. The serosal surface of the adjacent small bowel was abraded with a scalpel to produce punctuate bleeding. This area between the excised parietal peritoneum and adjacent small bowel serosa was then used for evaluating the efficacy of heparin for adhesion prevention. Alzet minipumps containing phosphate-buffered saline with varying doses of heparin were sewn into the right dorsal subcutaneous space with Vicryl sutures. A polyethylene catheter tip leading from the pump into the peritoneal cavity was placed 2-3 mm over the injury test site. Within seven days after the implantation of the pumps, the rabbits were killed and the extent of adhesions determined, according to a qualitative grading system. The threshold dosage at which significantly beneficial antiadhesion properties were obtained was 1.5 x 10(-2) U/hour or 7.5 x 10(-3) U/kg/day. Effective adhesion prevention occurred with only two days of treatment beginning the day of surgery. At this dose, no postsurgical bleeding occurred. These findings suggest that local intraperitoneal administration of low-dose heparin throughout the immediate postoperative interval may result in adhesion-free healing.

Acceleration of dermal tissue repair by angiotensin II.

Angiotensin II is a naturally occurring peptide which has been shown to possess angiogenic properties. In the studies reported here, angiotensin II was shown to increase the proliferation of cultured bovine aortic arch endothelial cells in a concentration-dependent manner. Acute administration of angiotensin II in Hydron accelerated the repair of dermal injuries in a full-thickness excisional rat model. Additional studies were done to determine the best vehicle for delivery of angiotensin II to a dermal injury. Several vehicles, including 10% low-viscosity carboxymethyl cellulose, 4% medium-viscosity carboxymethyl cellulose, and 3% high-viscosity carboxymethyl cellulose, were found to be effective in this regard. Daily administration of angiotensin II for days 0 to 4 after injury (day 0 being the time of surgery) was determined to provide the optimal dosage for acceleration of wound repair by angiotensin II. In addition, dose-response studies indicated that angiotensin II accelerated wound repair in a dose-dependent fashion with 0.03 and 0.01 microg/rat/day of angiotensin II administered on days 0 to 4 being the minimally effective and no-effect doses, respectively. Administration of 100 microg/day of angiotensin II in 10% carboxymethyl cellulose for 5 days after injury to animals with impaired healing (steroid- and adriamycin-treated rats and diabetic mice) was also found to accelerate the rate of repair. In conclusion, angiotensin II accelerated the closure of full-thickness skin injuries in a dose-dependent manner in normal and impaired animal models.

Intraperitoneal tolmetin prevents postsurgical adhesion formation in rabbits.

Previous studies showed that drugs that suppress prostaglandin synthesis were able to prevent adhesion formation following surgical trauma to the peritoneum. In our study, this premise was extended to examine the effects of various delivery systems on the ability of a nonsteriodal antiinflammatory drug, tolmetin, to prevent intraperitoneal adhesion formation after a standardized injury to the peritoneum. Continuous delivery of small amounts of tolmetin directly to the injury site following parietal peritoneal injury led to a significant reduction in the number and severity of adhesions formed. A single intraperitoneal injection of tolmetin in 5% Tween, however, required a relatively higher concentration of drugs to prevent adhesion formation. In a follow-up study of abrasion and devascularization of both uterine horns, the acid form of tolmetin was shown to be more potent than the sodium salt for the prevention of adhesion formation.

Effects of tolmetin sodium dihydrate on normal and postsurgical peritoneal cell function.

Recent studies utilizing intraperitoneal (i.p.) administration of NSAIDs to rabbits after surgical injury to the parietal peritoneum demonstrated macrophage involvement. NSAIDs are known to inhibit the metabolism of arachidonic acid to prostaglandins, which in turn modulate a variety of macrophage functions. Studies presented here examine the effects of tolmetin administration on peritoneal resident and post-surgical leukocyte functions, such as phagocytosis, the release of superoxide anion and tumoricidal activity. Rats, either non-surgical or following peritoneal surgery, were injected i.p. with tolmetin. At various times after treatment, the rats were sacrificed and peritoneal cells collected by lavage. The phagocytic capability of peritoneal leukocytes was transiently decreased 5-7 days after the administration of tolmetin to normal animals. However, administration of tolmetin during surgery extended the length of time that phagocytic capability was enhanced. In non-surgical controls, there was an elevation in superoxide anion release and tumoricidal activity 24 h after tolmetin administration. Superoxide anion release was suppressed at days 5 and 7 after treatment, but returned to control levels by day 14. Intraoperative administration of tolmetin significantly elevated superoxide anion release at days 3 and 5, phagocytosis at days 7 and 14 and tumoricidal activity at day 3. These studies suggest that compounds which suppress prostaglandin synthesis can modulate the function of resident and post-surgical peritoneal cells.