Comparison of ketorolac tromethamine with other injectable nonsteroidal anti-inflammatory drugs for pain-on-injection and muscle damage in the rat.

The local tolerance of ketorolac tromethamine (Toradol, Syntex) was compared with that of four other injectable nonsteroidal anti-inflammatory drugs (NSAIDs) (diclofenac sodium, piroxicam, ketoprofen, and metamizol magnesium) in the rat paw-lick/muscle irritation assay as described previously. All drugs were tested at concentrations approved for clinical use. After subplantar (footpad) injection, ketorolac produced virtually no pain-on-injection as assessed by the number of paw-lick/lift responses during a 15 min observation period. The other NSAIDs produced slight to moderate paw-lick/lift responses. Redness and swelling at the injection site were less severe for ketorolac than for the other NSAIDs. After intramuscular (i.m.) injection, all of the NSAIDs produced some degree of muscle damage, as assessed histopathologically 24 h after injection. The lesions, consisting primarily of muscle degeneration, were less severe for ketorolac than for the other NSAIDs. Ketorolac and metamizol produced the smallest elevations in serum creatine kinase, as measured 2 h after i.m. dosing, not significantly different from isotonic saline. Overall, ketorolac was better tolerated in the assay than the other injectable NSAIDs, thereby suggesting the possibility of improved local tolerance on clinical use.

Rat paw-lick/muscle irritation model for evaluating parenteral formulations for pain-on-injection and muscle damage.

A two-phase assay was developed in the rat to evaluate parenteral formulations intended for intramuscular administration for the induction of both acute pain-on-injection and delayed pain/discomfort at the injection site (secondary to muscle damage). Phase 1 of the assay assessed pain-on-injection using a modified version of the previously published rat paw-lick assay. Adult male CD rats (10/group) were given subplantar (footpad) injections of 0.1 ml and then observed for 15 min for paw-lick responses. To increase assay sensitivity, responses more subtle than paw licks (ie., paw lifts) were scored, and injection-site clinical signs were recorded 6, 24, and 48 hr after injection. Phase 2 of the assay assessed myotoxic potential, using the same rats after a 1-week recovery period. The rats were injected intramuscularly in the anterior thigh with 0.2 ml, bled from the orbital sinus at 2, 6, and 24 hr for analysis of serum creatine kinase (CK), and then necropsied at 24 hr to prepare tissue sections of the injection site for microscopic examination. A series of cephalosporin-type antibiotics produced pain-on-injection and muscle damage consistent with reported clinical experience (cefazolin less than cephalothin less than cefoxitin). Several nonantibiotic parenteral formulations (diazepam, digoxin, phenytoin, and lidocaine) tested in the paw-lick/muscle irritation model also produced responses that correlated with the clinic, i.e., virtually no acute pain but moderate to marked muscle damage. The results indicate that the two-phase rat paw-lick/muscle irritation model is effective in evaluating parenteral formulations for clinical acceptability, and that both phases of the assay are necessary to optimize predictability of the assay for human clinical experience.

Effect of polymyxin B on hemoglobin-mediated hepatotoxicity.

In previous studies, enhanced toxicity of hemoglobin when combined with endotoxin has suggested a specific binding site. This binding may take place even when endotoxin-free hemoglobin is administered, due to some low levels of endotoxin usually present in the portal vein. To test this hypothesis, we administered endotoxin-free, stroma-free human hemoglobin (SFH) to rabbits in doses shown previously to be hepatotoxic or lethal. A second group of rabbits received the anti-endotoxin antibiotic polymyxin B (PB) to inactivate endotoxin, followed by a dose of SFH. A control group received intravenous PB alone. There was no differences in the incidence of hepatotoxicity among these three groups. To show the binding interactions of endotoxin with PB and endotoxin with hemoglobin, an experiment using toluidine blue was performed. According to this study, specific binding of hemoglobin with endotoxin did not occur. Our results do not agree with previous suggestions that hemoglobin-mediated hepatotoxicity is due to the interaction in vivo of hemoglobin with low levels of endogenous endotoxin.

Respiratory bronchiolitis following long-term ozone exposure in bonnet monkeys: a morphometric study.

To quantitate the response of respiratory bronchiolar (RB) epithelium and peribronchiolar connective tissue (PCT) to chronic exposure to high ambient levels of ozone, two groups of 8 adult male bonnet monkeys each were subjected 8 h daily for one year to 0.64 ppm (UV standard) ozone or filtered air, respectively. Blocks of tissue selected throughout the lung and from first generation RBs following airway microdissection had the following significant exposure-related changes: 57% greater volume of RB in the lung, 27% smaller diameter of RB lumen, 179% thicker media and intima of peribronchiolar arterioles, 61% thicker RB epithelium, and 77% thicker PCT. The increase in thickness of the RB wall resulted primarily from an 84% increase in PCT, with the remainder from the epithelium. Estimates of cellular numerical density showed an 81% increase in cuboidal bronchiolar cells and an 87% decrease in type 1 pneumocytes in the exposed group. Cell volumes from serial section reconstruction showed significantly larger cuboidal bronchiolar (79%), ciliated (117%), and type 2 (66%) cells over controls. Significant PCT changes included more amorphous extracellular matrix (288%), neutrophils (1523%), and lymphocytes/plasma cells (307%). The number of fibroblasts and the volume of extracellular fibers were larger than control values by 44% and 31% in the exposed group, but these changes were not statistically significant. Centriacinar changes due to exposure to long-term, high ambient ozone in bonnet monkeys results in narrowing of respiratory bronchioles primarily by peribronchiolar inflammation (inflammatory cells, fibers, amorphous matrix) and secondarily through hyperplasia of cuboidal bronchiolar cells.

Structure as revealed by airway dissection. A comparison of mammalian lungs.

Microdissection of mammalian pulmonary airways demonstrates branching patterns and provides precisely defined tissue samples for morphologic study. The dissections are performed on lung fixed by airway infusion at standard pressures. Using fine scissors and a high resolution dual-viewing dissecting microscope, extrapulmonary and intrapulmonary airways are dissected down their axial pathways. The plane of dissection is chosen to include as many minor daughter (side) branches as possible. Lungs from 5 species: sheep, goat, cat, rabbit, and bonnet monkey have been dissected, photographed, successive generations numbered, and pieces of tissue processed for LM, TEM, and SEM. Branching patterns differ between lobes (cranial versus caudal) of the same species and between the same lobe in different species. Marked differences in epithelial population distribution within the airway tree are found between the same lobe of different species (i.e., cranial lobes of rabbit and sheep) and between different lobes in the same species (i.e., cranial and caudal lobes of the sheep). The dissection approach to pulmonary airway morphologic studies provides specimens of precisely defined branching history, generation number, and anatomic position within regions of the lung and within specific segments. This allows studies that compare: (1) different airway generations in the same pathway, (2) bifurcation points and the airway segments between them, (3) terminal airways of differing pathway lengths and numbers of branching, (4) terminal airways of different regions of same lobe, (5) same airway generations in different lobes, and (6) same airway generations from animal to animal and species to species.

Extrahepatic biliary atresia in a rhesus monkey (Macaca mulatta).

Extrahepatic biliary atresia was observed in a 6-week-old female rhesus monkey. Jaundice and conjugated hyperbilirubinemia were detected at the age of 6 days and persisted throughout life. At 6 weeks of age, the diagnosis of extrahepatic biliary atresia was established at exploratory laparotomy, and bile duct remnants were biopsied. Histological examination of these specimens showed inflammatory and fibrosing lesions similar to those observed in humans with extrahepatic biliary atresia. Because of serologic evidence of Reovirus 3 infection in human patients with extrahepatic biliary atresia, serum of the affected monkey was tested for antibodies to this virus. Three sequential serum samples obtained during the course of illness showed persistently high Reovirus 3 titers which are consistent with but do not prove concurrent Reovirus 3 infection. This report represents the first documented case of spontaneous extrahepatic biliary atresia in a nonhuman primate and suggests that this species may be suitable for further investigation of the pathogenesis of this disease.

Work-induced muscle hypertrophy in vitamin D-deficient rats.

To determine if vitamin D deficiency would retard the ability of muscle to hypertrophy in response to mechanical stress, we severed the gastrocnemius tendon on one leg of rats in each of three groups, the treatment of which differed only in the amount of vitamin D in the diet. After 1 week the increased size of the soleus and plantaris in the leg in which the gastrocnemius was severed relative to that of the sham operated leg, was determined for each rat. Despite differences in body weight and serum calcium among the groups, we found no difference in the percent of muscle hypertrophy. We conclude that muscle hypertrophy can occur in response to local mechanical forces despite a deficient hormonal environment that otherwise retards growth.

Changes in the manufacture of expanded microporous polytetrafluoroethylene: effects on patency and histological behavior when used to replace the superior vena cava.

Polytetrafluoroethylene (PTFE) has been used clinically and experimentally as a vein substitute. Since its introduction, changes have been made in the manufacture of the material. This study was designed to evaluate the effects of pore size (fibril length), PTFE cover, and spiral tube support on long-term patency and histological behavior when this material is used as a replacement for the precava in the dog. A graft of 30-mu pore size has adequate fibroblastic ingrowth, a neointima of 200 mu or less, and the best long-term patency. The PTFE cover results in less fibroblastic involvement of the prosthetic graft and prevents late occlusion caused by transmural fibrosis in the 90-mu graft. The spiral support does not affect patency and may be indicated when external pressure on the tube must be avoided.