Application of a simple measuring method to evaluate the fecal microflora of dairy cows in the summer season.

OBJECTIVE: The effect of seasonality needs to be considered in designing future studies because global warming has caused a rise in ambient temperatures. The objective of the present study is to investigate the effect of high ambient temperatures on fecal score and fecal microflora in dairy cows during summer. MATERIALS AND METHODS: During the 7 days before the sampling of feces, the daily mean temperatures were 19.9°C in early summer and more than 27.5°C in late summer. Fecal samples were collected from the rectum of cows and the fecal score was evaluated on a 4-point scale. The equalized samples were used to extract the genomic deoxyribonucleic acid (DNA) of the bacteria (Escherichia coli, Salmonella, Lactobacillus, and Bifidobacterium). RESULTS: There was no significant difference in fecal scores between the sampling times in early and late summer. In the populations of the bacteria, there was no significant difference between sampling days in the DNA level of Salmonella, and E. coli in late summer increased to more than three times the level in early summer. However, both levels of Lactobacillus and Bifidobacterium in early summer significantly decreased after 2 months. CONCLUSION: These data suggest that the increase in temperature in late summer may adversely affect the populations of bacteria in the intestinal environment of dairy cows. In addition, the method used in the present study was sufficient to evaluate the changes in internal and external environmental conditions of dairy cattle.

Effects of Simvastatin on Alveolar Regeneration and Its Relationship to Exposure in Mice with Dexamethasone-Induced Emphysema.

In the present study, the relationship between systemic exposure of simvastatin (SV) hydroxy acid (SV-acid), an active form of SV, and its alveolar regeneration rates was investigated using emphysema model mice created by postnatal treatment of dexamethasone. In a model with young animals, the mice were treated with SV for 10 d from postnatal day 42. Similar alveolar regeneration with a % mean linear intercept (Lm) recovery of 60 to 70% by histochemical observation was observed in mice after intraperitoneal administration at dose in the range of 4-100 µg/mouse. The % Lm recovery after oral administration of 20 µg/mouse was comparable with that after intraperitoneal administration at a dose of 4 µg/mouse, when their exposure of SV-acid was almost similar in both treated groups. Regardless of the route of administration, the recovery can depend on the exposure level of SV-acid, and to the maximum was about 60-70%. On the other hand, in a model with adult animals, the mice were intraperitoneally administrated SV at a dose of 4 µg/mouse for 10 d from postnatal day 152. Compared to young animals, less % Lm recovery was observed in adult mice even their systemic exposures of SV-acid were similar.

Pharmacokinetics of ivermectin applied topically by whole-body bathing method in healthy volunteers.

As a novel administration method of ivermectin (IVM) for scabies treatment, we proposed a "whole-body bathing method (WBBM)". In this method, the patients would bathe themselves in a bathing fluid containing IVM at an effective concentration. Previously, we demonstrated that WBBM could deliver IVM to the skin but not to the plasma in rats. In the present study, to assess the clinical validity of the method an arm bathing examination (first trial) and a whole-body bathing examination (second trial) were conducted in healthy volunteers. In both the first and second trials, after bathing in fluid containing IVM, the exposure in the stratum corneum was higher compared with that after taking IVM p.o. as reported previously. IVM was not detected in plasma at any sampling point after the whole-body bathing in the second trial. Furthermore no serious adverse events were found. These results in both trials suggest that WBBM can deliver IVM to the human stratum corneum without systemic exposure or serious adverse effects in healthy volunteers, and at concentrations that would be adequate for scabies treatment.

Bioavailability of an R-α-Lipoic Acid/γ-Cyclodextrin Complex in Healthy Volunteers.

R-α-lipoic acid (R-LA) is a cofactor of mitochondrial enzymes and a very strong antioxidant. R-LA is available as a functional food ingredient but is unstable against heat or acid. Stabilized R-LA was prepared through complexation with γ-cyclodextrin (CD), yielding R-LA/CD. R-LA/CD was orally administered to six healthy volunteers and showed higher plasma levels with an area under the plasma concentration-time curve that was 2.5 times higher than that after oral administration of non-complexed R-LA, although the time to reach the maximum plasma concentration and half-life did not differ. Furthermore, the plasma glucose level after a single oral administration of R-LA/CD or R-LA was not affected and no side effects were observed. These results indicate that R-LA/CD could be easily absorbed in the intestine. In conclusion, γ-CD complexation is a promising technology for delivering functional but unstable ingredients like R-LA.

Effects of All trans-Retinoic Acid on Alveolar Regeneration in Dexamethasone-Induced Emphysema Models and Its Relationship to Exposure in ICR and FVB Mice.

During the past two decades, it has been reported that treatment with all-trans-retinoic acid (ATRA) induces alveolar regeneration in rodent emphysema models. In the present study, we investigated the regeneration by ATRA at various exposure conditions in two strains of mice with induced emphysema. The emphysema model was created by postnatal administration of dexamethasone to ICR and FVB mice, which were then treated with ATRA from postnatal day 42. The regeneration was observed in ICR mice but not in FVB mice given 10 and 40 mg/kg/d ATRA for 10 d. The concentration-time profiles of ATRA in plasma and lung were similar in both strains. These results suggest that the strain difference in the regeneration by ATRA was not caused by differences in the exposure to ATRA. On the other hand, the regeneration in ICR mice was enhanced by an increase of the intraperitoneal dose in the range of 10-40 mg/kg/d for 10 d. At an intraperitoneal dose of 40 mg/kg/d, the regeneration was observed after 10 and 20 d of treatment but not after 1 to 5 d of treatment. Meanwhile, the regeneration by intraperitoneal administration of ATRA was enhanced more than that by oral administration. Exposure to ATRA during repeated intraperitoneal administration to ICR mice was higher than that in oral administration. The results suggest that the regeneration in ICR mice requires at least 10 d of treatment with ATRA and its effects depend on the exposure to ATRA in plasma, which parallels that in lung.

Effect of tiopronin on excretion of homocysteine and its plasma exposure level in rats.

When the biosynthesis of homocysteine (Hcy), an amino acid containing thiol, exceeds its elimination, plasma Hcy concentration increases and results in hyperhomocysteinemia (HHC). Most of the Hcy in plasma covalently binds to the cysteine residues of albumin by disulfide bonds. Meanwhile tiopronin (TP), a thiol-containing compound, is used for treatment of cysteinuria to decrease the amount of insoluble cystine in urine, by replacing a cysteine molecule in cystine with TP to form soluble TP-cysteine complexes. The present study investigated the effect of TP on the total (protein-unbound and protein-bound) Hcy concentration and the ratio of protein-unbound Hcy to total Hcy in plasma, and the urinary excretion of Hcy. Methionine was administered orally to rats to induce temporary hyperhomocysteinemia, and then TP was administered and the plasma and urine were collected. The amount of Hcy excreted in urine was higher but the plasma concentration of Hcy was lower in the TP group than in the control group until 3 h after TP administration. In addition the ratio of protein-unbound Hcy in plasma tended to be increased by TP administration. These results demonstrated that TP enhanced the urinary excretion of Hcy, which might cause the decrease of its plasma concentration in rats.

Effect of high-fat meal intake on the pharmacokinetic profile of ivermectin in Japanese patients with scabies.

Ivermectin (IVM) is used as an anthelmintic agent in many countries. To evaluate the effect of high-fat (HF) meal intake on the pharmacokinetics of IVM, a clinical trial was conducted in Japanese patients with scabies. The patients were administrated Stromectol(®) tablets in the fasted state, and after 1 week they were also administrated it after a HF meal (fed state). After the administration, IVM concentrations in plasma and the stratum corneum were determined. The geometric mean of fed/fasted ratio of area under IVM concentration-time curve (AUC) in plasma was 1.25 (90% confidence interval, 1.09-1.43), suggesting the tendency to increased absorption after a HF meal. The fed/fasted ratio of the maximum IVM concentration in the stratum corneum was well correlated with that in plasma. In addition, no serious adverse events were observed during the trial, while a mild increase of aspartate aminotransferase and alanine aminotransferase activity in plasma was observed under the fed state in two patients. The mean AUC of IVM in plasma of those two patients were approximately threefold higher than that of the other patients at that time. On the other hand, the treatment success rate was 76.9% at 7 days after the second administration, which was comparable with the expected level. The present study not only demonstrates that HF meal intake increases the IVM concentration in plasma and the stratum corneum in Japanese patients with scabies, but also suggests the possibility that HF meals increase the risk of hepatic dysfunction by the increased exposure of IVM.

Investigation of Enantioselective Membrane Permeability of α-Lipoic Acid in Caco-2 and MDCKII Cell.

α-Lipoic acid (LA) contains a chiral carbon and exists as two enantiomers (R-α-lipoic acid (RLA) and S-α-lipoic acid (SLA)). We previously demonstrated that oral bioavailability of RLA is better than that of SLA. This difference arose from the fraction absorbed multiplied by gastrointestinal availability (F(a) × F(g)) and hepatic availability (F(h)) in the absorption phase. However, it remains unclear whether F(a) and/or F(g) are involved in enantioselectivity. In this study, Caco-2 cells and Madin-Darby canine kidney strain II cells were used to assess the enantioselectivity of membrane permeability. LA was actively transported from the apical side to basal side, regardless of the differences in its steric structure. Permeability rates were proportionally increased in the range of 10-250 µg LA/mL, and the permeability coefficient did not differ significantly between enantiomers. Hence, we conclude that enantioselective pharmacokinetics arose from the metabolism (F(h) or F(g) × F(h)), and definitely not from the membrane permeation (F(a)) in the absorption phase.

Evaluation of cytochrome P450 inductions by anti-epileptic drug oxcarbazepine, 10-hydroxyoxcarbazepine, and carbamazepine using human hepatocytes and HepaRG cells.

Anti-epileptic drug oxcarbazepine is structurally related to carbamazepine, but has reportedly different metabolic pathway. Auto-induction potentials of oxcarbazepine, its pharmacologically active metabolite 10-hydroxyoxcarbazepine and carbamazepine were evaluated by cytochrome P450 (CYP) 1A2, CYP2B6 and CYP3A4 mRNA levels and primary metabolic rates using human hepatocytes and HepaRG cells. For the CYP1A2 the induction potential determined as the fold change in mRNA levels was 7.2 (range: 2.3-11.5) and 10.0 (6.2-13.7) for oxcarbazepine and carbamazepine, respectively, while 10-hydroxyoxcarbazepine did not induce. The fold change in mRNA levels for CYP2B6 was 11.5 (3.2-19.3), 7.0 (2.5-10.8) and 14.8 (3.1-29.1) for oxcarbazepine, 10-hydroxyoxcarbazepine and carbamazepine, respectively. The fold change for CYP3A4 induction level by oxcarbazepine, 10-hydroxyoxcarbazepine and carbamazepine was 3.5 (1.2-7.4), 2.7 (0.8-5.7) and 8.3 (3.5-14.5), respectively. The data suggest lower induction potential of oxcarbazepine and 10-hydroxyoxcarbazepine relative to carbamazepine. The results in HepaRG cells showed similar trend as the human hepatocytes. After incubation for 72 h in hepatocytes and HepaRG cells, auto-induction was evident for only carbamazepine metabolism. The 10-keto group instead of double bond at C10 position is evidently a determinant factor for limited auto-induction of P450 enzymes by oxcarbazepine.

Mechanism for transport of ivermectin to the stratum corneum in rats.

Ivermectin (IVM) is used as an oral medication for scabies, a skin infection caused by a mite, sarcoptes scabiei, which parasitizes in the stratum corneum. After oral administration IVM is absorbed from the intestine, and finally distributed to the stratum corneum to eliminate the mites. However its transport mechanism remains unclear. A pharmacokinetic study was performed using hairless Wistar Yagi (HWY) rats, which have no or atrophied sebaceous glands, and Wistar rats as a reference. After oral administration of IVM to both groups, the area under the concentration-time curve of IVM in the dermis and epidermis (dermis-epidermis) of HWY rats were about 60% lower than that of Wistar rats, even though the plasma concentration profiles were comparable in both groups. In addition at 12 h after the administration, IVM concentration in the outer stratum corneum, the shallower layer of the dermis-epidermis, was higher compared to that in the deeper layer. In the dermis-epidermis of the skin from various locations, the concentrations of IVM and squalene, the latter of which is secreted to the skin surface via the sebaceous gland, were positively well correlated. Those results suggest that IVM is transported to the stratum corneum via the sebaceous glands.

Enantioselective Pharmacokinetics of α-Lipoic Acid in Rats.

α-Lipoic acid (LA) is widely used for nutritional supplements as a racemic mixture, even though the R enantiomer is biologically active. After oral administration of the racemic mixture (R-α-lipoic acid (RLA) and S-α-lipoic acid (SLA) mixed at the ratio of 50:50) to rats, RLA showed higher plasma concentration than SLA, and its area under the plasma concentration-time curve from time zero to the last (AUC) was significantly about 1.26 times higher than that of SLA. However, after intravenous administration of the racemic mixture, the pharmacokinetic profiles, initial concentration (C0), AUC, and half-life (T1/2) of the enantiomers were not significantly different. After oral and intraduodenal administration of the racemic mixture to pyrolus-ligated rats, the AUCs of RLA were significantly about 1.24 and 1.32 times higher than that of SLA, respectively. In addition, after intraportal administration the AUC of RLA was significantly 1.16 times higher than that of SLA. In conclusion, the enantioselective pharmacokinetics of LA in rats arose from the fraction absorbed multiplied by gastrointestinal availability (FaFg) and hepatic availability (Fh), and not from the total clearance.

Population pharmacokinetic analysis for 10-monohydroxy derivative of oxcarbazepine in pediatric epileptic patients shows no difference between Japanese and other ethnicities.

Oxcarbazepine is an anti-epileptic drug, which is almost completely metabolized by cytosolic enzymes in the liver to the active 10-monohyroxy metabolite (MHD) following oral administration. The pharmacokinetic (PK) profiles of MHD were evaluated in pediatric epileptic patients and a possible ethnic difference in PK of MHD between Japanese and non-Japanese pediatric patients was assessed. A non-linear mixed effect modeling approach was used to determine the PK of MHD. A one-compartment population model with first-order absorption appropriately described the PK of MHD. No clinically relevant differences were found for using body surface area or weight to explain between-patient variability, therefore the final model included the effects of body weight on apparent clearance (CL/F) and apparent volume of distribution (V/F) of MHD, and in addition, the effect of 3 concomitant anti-epileptic drugs (carbamazepine, phenobarbital and phenytoin) on CL/F of MHD. Inclusion of ethnicity as a covariate in the final model, concluded no ethnic difference with respect to CL/F of MHD between Japanese and non-Japanese patients. Hence, oxcarbazepine can be generally applied using the same dosage and administration for the treatment of partial onset seizures in pediatric patients, regardless of ethnicity.

Effect of γ-Cyclodextrin Inclusion Complex on the Absorption of R-α-Lipoic Acid in Rats.

R-α-lipoic acid (RLA) is an endogenous organic acid, and works as a cofactor for mitochondrial enzymes and as a kind of antioxidant. Inclusion complexes of RLA with α-, ß- or γ-cyclodextrins (CD) were prepared and orally administered as a suspension to rats. Among them, RLA/γ-CD showed the highest plasma exposure, and its area under the plasma concentration-time curve (AUC) of RLA was 2.2 times higher than that after oral administration of non-inclusion RLA. On the other hand, the AUC after oral administration of non-inclusion RLA and RLA/γ-CD to pylorus-ligated rats did not differ. However, the AUC after intraduodenal administration of RLA/γ-CD was 5.1 times higher than that of non-inclusion RLA, and was almost comparable to the AUC after intraduodenal administration of RLA-Na solution. Furthermore, the AUC after intraduodenal administration of RLA/γ-CD was not affected by biliary ligation or co-administration of an amylase inhibitor. These findings demonstrated that RLA was absorbed from the small intestine effectively when orally administered as a γ-CD inclusion complex, which could be easily dissolved in the lumen of the intestine. In conclusion, γ-CD inclusion complex is an appropriate formulation for supplying RLA as a drug or nutritional supplement with respect to absorption.

Effect of high fat intake on the pharmacokinetic profile of ivermectin in rabbits.

Ivermectin (IVM) is used as an oral drug for treatment of scabies. It was reported that the area under the plasma concentration-time curve (AUC) of IVM was higher in healthy volunteers after a high-fat meal than in those who were fasting, but the mechanism has not been clarified yet. In fasted rabbits, the AUC after oral administration of IVM as a solution was higher than that of a suspension, indicating that the absorption of IVM depends on how much is dissolved in the gastrointestinal tract. On the other hand, the AUC was higher in rabbits pre-treated with a high-fat solution (HF; fat and cholesterol) than in those that had fasted, when IVM was administered orally not only in suspension but also in solution, and even when it was administered intravenously. In addition, the increase in total cholesterol level in the HF condition was correlated with the increase in IVM level. These results suggest that enhancement of solubility may not be the only reason for the increase of AUC in rabbits. It is also suggested that the increase of cholesterol concentration might change the distribution profile of IVM in plasma, and accordingly its concentration could be increased.

Dysfunction of blood pressure regulation in hyperhomocyteinemia model in rats.

Hyperhomocysteinemia (HHC) has been reported to be one of risk factors for cardiovascular disease. We investigated the effects of HHC on blood pressure regulation and its association with damage to the thoracic aorta and imbalance of redox in plasma in rats. Rats were fed a methionine enriched diet (Met diet) or a methionine and cholesterol (Met+Chol diet) enriched diet for 16 weeks to create a subchronic HHC model, in which the plasma concentration of homocysteine was about 7 times higher than that of control rats. The increase in systolic blood pressure (Δ-SBP) from sympathetic stimulation by L-epinephrine was 2- to 3-fold larger in HHC model in rats than that in control rats after several weeks of the treatment. These findings suggest that HHC deteriorates vaso-regulatory function, which could bring on an increased risk of cardiovascular events in humans. In addition, some of the elastic lamellae in the aorta were disrupted in the HHC group. However, the content of cross-linkages which give elasticity and mechanical strength in the lamellae, was not significantly different between HHC and control rats. Also plasma concentrations of thiobarbituric acid reactive substance and glutathione as indicators for redox balance in plasma were not different. In conclusion, the deterioration of vaso-regulation in HHC model in rats might be caused by the damage to elastic lamellae in the aorta, and not by oxidative stresses.

Experimental study of pharmacokinetics of external, whole-body bathing application of ivermectin.

As a novel method improving the safety of conventional oral ivermectin (IVM) for scabies treatment, we conceived an idea called the "whole-body bathing method". In this method, the patients would bathe themselves in a bathing fluid containing IVM at an effective concentration. To evaluate the feasibility of the method, we investigated the IVM concentration in the skin and plasma after bathing rats in a fluid containing 100 ng/mL of IVM. After the bathing, the concentration of IVM in the skin was more than 400 ng/g wet weight and was maintained until 8 h after the bathing. The concentration was clearly higher than that in patients taking IVM p.o. as previously reported; IVM was not detected in plasma in the present study. Thus, the method would be a preferable drug delivery system for the skin application of IVM compared with p.o. administration.

Serum concentrations of clarithromycin and rifampicin in pulmonary Mycobacterium avium complex disease: long-term changes due to drug interactions and their association with clinical outcomes.

BACKGROUND: Concomitant use of clarithromycin (CAM) and rifampicin (RFP) for the treatment of pulmonary Mycobacterium avium complex (MAC) disease affects the systemic concentrations of both drugs due to CYP3A4-related interactions. To date, however, there has been no report that investigates the long-term relationship between the drug concentrations, CYP3A4 activity, and clinical outcomes. Our aim was to investigate the time course of the drug levels in long-term treatment of subjects with pulmonary MAC disease, and examine the correlation of these concentrations with CYP3A4 activity and clinical outcomes. METHODS: Urine and blood samples from nine outpatients with pulmonary MAC disease were collected on days 1, 15, and 29 (for four subjects, sample collections were continued on days 57, 85, 113, 141, 169, 225, 281, 337, and 365). Serum drug concentrations and urinary levels of endogenous cortisol (F) and 6 beta-hydroxycortisol (6ßOHF), the metabolite of F by CYP3A4, were measured, and evaluated 6ßOHF/F ratio as a CYP3A4 activity marker. In addition, the clinical outcomes of 4 subjects were evaluated based on examination of sputum cultures and chest images. RESULTS: The mean 6ßOHF/F ratio increased from 2.63 ± 0.85 (n = 9) on the first day to 6.96 ± 1.35 on day 15 and maintained a level more than double initial value thereafter. The serum CAM concentration decreased dramatically from an initial 2.28 ± 0.61 µg/mL to 0.73 ± 0.23 µg/mL on day 15. In contrast, the serum concentration of 14-hydroxy-CAM (M-5), the major metabolite of CAM, increased 2.4-fold by day 15. Thereafter, both CAM and M-5 concentrations remained constant until day 365. The explanation for the low levels of serum CAM in pulmonary MAC disease patients is that RFP-mediated CYP3A4 induction reached a maximum by day 15 and remained high thereafter. Sputum cultures of three of four subjects converted to negative, but relapse occurred in all three cases. CONCLUSIONS: Our study demonstrated that serum CAM concentrations in pulmonary MAC disease patients were continuously low because of RFP-mediated CYP3A4 induction, which may be responsible for the unsatisfactory clinical outcomes.

Effects of postnatal dexamethasone treatment on mRNA expression profiles of genes related to alveolar development in an emphysema model in mice.

Emphysema can be induced in animals by postnatal treatment with dexamethasone (Dex) and such models have been widely used for various research. However, it is not clear what are the effects of Dex on assembly of alveolar elastic fibers in the emphysema model in mice. This study compared the expression profile of genes related to alveolar development between Dex treated and control mice during the treatment from postnatal day 3 (P3) to P14 with a 2-day break. From morphological observation of lung sections on P42, we confirmed the induction of emphysema in the treated mice. The mRNA expression level of fibrillin-1, which consists of microfibrils as a scaffold to form elastic fibers, and fibulin-5, which is a key protein reinforcing the fibers, reached maximum on P7 in control mice. However, in the Dex group, expression levels both types of mRNA were much lower with no clear expression peak. On the other hand, mRNA expression of tropoelastin, the main component in elastic fibers, reached maximum on P5 in the Dex group, which was 9 days earlier than in the control group. At this time, the amount of microfibrils might not be enough for tropoelastin to be deposited completely in Dex treated mice. This imbalance in the expression of tropoelastin and microfibril might interfere with the efficient formation of elastic fibers.

Disruption of elastic lamellae in the aorta by D-penicillamine and its effect on vaso-regulation in rats.

We assessed the effects of D-penicillamine (D-PA) on cross-linkages in elastin and vaso-regulatory function in rats. After administration of D-PA at a dose of 100 mg/kg/day for 7 weeks to adult and young rats, the thoracic aortas were isolated. The elastic lamellae in the aorta were disrupted histopathologically in all the treated groups. The content of cross-linkages in elastin, i.e. desmosine and isodesmosine, which gives elasticity to the aortic wall, was significantly reduced in the D-PA treated groups versus the control groups. On the other hand, the content of pyridinoline as a marker of insoluble collagen was significantly reduced in the D-PA treated groups, even though the total collagen content was not changed. In addition, after 7 weeks of treatment with D-PA, the change between systolic blood pressure before and after sympathetic stimulation (Δ-SBP) by L-epinephrine was about 2.5-fold larger than that in the control group. Similar results were obtained using angiotensin II or ouabain instead of L-epinephrine. These findings demonstrated that D-PA disrupted elastic lamellae of the rat aorta by reduction of the cross-linkages in elastin and collagen, which caused dysfunction of vaso-regulation. Also, they suggested the possibility that long-term treatment with D-PA in patients could cause a decrease in vaso-regulatory function and could increase the risk of cardiovascular events.

Disruption of elastic lamellae in aorta and dysfunction of vaso-regulation by rofecoxib in rats.

We assessed the effects of rofecoxib on cross-linkage formation in elastin and vaso-regulatory function in rats. After administration of rofecoxib at a dose of 10 mg/kg for 7 weeks to young rats and for 7 and 10 weeks to adult rats, thoracic aortas were isolated. The elastic lamellae in the aortas were disrupted histopathologically in all the treated groups. However, the content of cross-linkages in elastin, i.e. desmosine and isodesmosine, which give elasticity to the aortic wall, was not significantly different between the rofecoxib treated and control groups. On the other hand, although the baseline blood pressure was not changed during the treatment period in both young and adult rats, after several weeks of treatment with rofecoxib the change between systolic blood pressure before and after sympathetic stimulation by L-epinephrine was 2 to 3-fold larger than that in the control group. Similar results were obtained using angiotensin II instead of L-epinephrine. The exposure to rofecoxib (area under the plasma concentration-time curve) of rats after single administration was a few times higher than that of humans in clinical use. These findings indicate that rofecoxib did not directly inhibit formation of cross-linkages in elastin of the aorta in rats. However, the treatment with rofecoxib for several weeks disrupted elastic lamellae and caused depression of vaso-regulatory function in rats, which could bring on an increased risk of cardiovascular events in human.