Groups IM Research Articles

Effect of Acute Administration of Ethanol Extract of Cannabis sativa Leaf on Oxidative Stress Biomarkers in Male and Female Wistar Rats

Oxidative stress is a phenomenon caused by an imbalance between production and accumulation of oxygen reactive species (ROS) in the cells and tissues and the ability of a biological system to detoxify these reactive products. This study investigated the acute effect of different doses of ethanolic extract annabis sativa (EECS) on oxidative stress biomarkers in male (M) and female (F) Wister rats. Twenty male rats (170 g ± 1.24) and twenty female rats (150 g ± 1.05) were separately assigned into four groups of five animals each for male and female, such that the rats in groups IM and IF, IIM and IIF, IIIM and IIIF and IVM and IVF received orally 1 mL of distilled water, 2 mg/kg body weight (BW) of EECS, 4 mg/kg BW of EECS and 6 mg/kg BW of EECS respectively for twenty one (21) days. Catalase, superoxide dismutase (SOD), Glutathione peroxidase (GPx), Glutathione reductase (GSH), malondialdehyde (MDA), total antioxidant capacity (TAC) and lactate dehydrogenase (LDH) were determined using standard methods. Administration of different doses (2 mg/kg BW, 4 mg/kg BW and 6 mg/kg BW) of EECS significantly (p<0.05) decreased catalase, SOD, GPx, GSH and TAC levels respectively when compared with the control. However, Cannabis sativa (CS) increased MDA and LDH levels significantly (p<0.05) when compared with the control. It was deduced that these alterations in oxidative stress biomarkers were dependent on the doses of CS consumed. Thus, the more the concentration of CS consumed, the more it may affect oxidative stress biomarkers negatively. In addition, all these effects of CS on oxidative stress biomarkers were more in male than in female.

Doxycycline Attenuates Doxorubicin-Induced Cardiotoxicity by Improving Myocardial Energy Metabolism in Rats.

Aim: Evaluate the influence of doxycycline, an anti-inflammatory and matrix metalloproteinase (MMP) inhibitor, on the attenuation of chronic doxorubicin-induced cardiotoxicity in rats. Methods: We allocated male Wistar rats into four groups: control (C), doxorubicin (D), doxycycline (inhibitor of MMP, IM), and Dox + doxycycline (DIM). Groups IM and DIM received doxycycline (5 mg/kg, IP) once a week for 4 weeks. In addition, 48 h after every doxycycline injection, groups D and DIM received Dox (5 mg/kg, IP). We performed echocardiogram and evaluated TIMP-4 and collagen I protein expression, MMP-2 activity, and oxidative stress and myocardial metabolism. Results: Doxorubicin promotes left atrium (LA) and left ventricle (LV) dilatation and decreases in LV fractional shortening, which was improved by doxycycline. Moreover, doxycycline attenuated the LV cardiomyocyte hypertrophy and collagen type I expression. Doxorubicin increased phosphofructokinase and decreased beta-hydroxyacyl Co-A dehydrogenase, pyruvate dehydrogenase, citrate synthase, and ATP synthase activity, which was partially attenuated by doxycycline. Lastly, doxycycline improved antioxidant enzyme activity in the DIM group. Conclusion: Doxorubicin increases oxidative stress and promotes changes in myocardial energy metabolism, accompanied by structural and functional changes. Doxycycline attenuated the doxorubicin-induced cardiotoxicity, at least in part, through changes in myocardial energy metabolism.

Comparative immunohistochemical assessment of the effect of repetitive anesthesia with isoflurane and sevoflurane on rat liver

Inhalation anesthetics are used in human, as well as veterinary medical practice. In the present study we investigated the effect of isoflurane and sevoflurane on rat hepatocytes. A total of 40 Wistar female rats were used in this study. Animals were divided in groups of 5 rats. Groups IM, SM served as control groups. Groups I1, I2, I3 were used to study isoflurane and S1, S2, S3 for sevoflurane study. They were anesthetized 3 times, for 2h long, at 2 days interval with a concentration of: 1.5% isoflurane (I1, I2, I3) and 2% sevoflurane (S1, S2, S3). The oxygen supply throughout the anesthesia was 1LO2/min. Groups IM, IS, I1, S1 were sacrificed immediately after the last anesthesia. Groups I2, S2 were sacrificed 6h after the last anesthesia, and groups I3, S3, 24h post-anesthesia. Liver samples were harvested to highlight caspase-3 in apoptotic hepatocytes. Following isoflurane administration, there were less than 1% cells in apoptosis highlighted in rat livers from groups IM, I1 and I2. At 24h post-anesthesia (group I3), a small number of apoptotic hepatocytes was highlighted (around 3.23% cells in apoptosis), with a strictly periacinar disposition, randomly distributed in a small number of hepatic lobules. After sevoflurane administration, less than 1% apoptotic hepatocytes were identified at all control moments throughout the study. The results suggest that the anesthetics do not present a considerable hepatotoxicity. The comparative assessment of the two anesthetics shows that sevoflurane is superior to isoflurane.

Comparative immunohistochemical assessment of the effect of repetitive anesthesia with isoflurane and sevoflurane on rat liver

Background and objectivesInhalation anesthetics are used in human, as well as veterinary medical practice. In the present study we investigated the effect of isoflurane and sevoflurane on rat hepatocytes. MethodsA total of 40 Wistar female rats were used in this study. Animals were divided in groups of 5 rats. Groups IM, SM served as control groups. Groups I1, I2, I3 were used to study isoflurane and S1, S2, S3 for sevoflurane study. They were anesthetized 3 times, for 2h long, at 2 days interval with a concentration of: 1.5% isoflurane (I1, I2, I3) and 2% sevoflurane (S1, S2, S3). The oxygen supply throughout the anesthesia was 1LO2/min. Groups IM, IS, I1, S1 were sacrificed immediately after the last anesthesia. Groups I2, S2 were sacrificed 6h after the last anesthesia, and groups I3, S3, 24h post-anesthesia. Liver samples were harvested to highlight caspase-3 in apoptotic hepatocytes. ResultsFollowing isoflurane administration, there were less than 1% cells in apoptosis highlighted in rat livers from groups IM, I1 and I2. At 24h post-anesthesia (group I3), a small number of apoptotic hepatocytes was highlighted (around 3.23% cells in apoptosis), with a strictly periacinar disposition, randomly distributed in a small number of hepatic lobules. After sevoflurane administration, less than 1% apoptotic hepatocytes were identified at all control moments throughout the study. ConclusionsThe results suggest that the anesthetics do not present a considerable hepatotoxicity. The comparative assessment of the two anesthetics shows that sevoflurane is superior to isoflurane.

Oxidative Stress Evaluation in Rats Anesthetized with Isoflurane or Sevoflurane

Oxidative stress implies an imbalance between antioxidant production and reactive oxygen species generation. In this experimental study, we aimed to assess the effects of two inhalational anesthetics (isoflurane and sevoflurane) on the oxidative stress enzymes, namely: superoxid dismutase and glutathione peroxidase. Our protocol included 40 Wistar female rats, divided into 8 groups (5 animals per group): IM, I1, I2, I3, SM, S1, S2 and S3. The groups were subdued to: oxygen alone (control groups IM and SM), isoflurane (I1, I2, I3) and sevoflurane anesthesia (S1, S2, S3), 3 times, for 2 hours long, every other day. The supplied oxygen was 1l O 2 /min and the concentration of isoflurane and sevoflurane was 1 minimum alveolar concentration (1 MAC). We harvested blood samples from each group at different moments: IM, SM, I1, S1 - immediately, I2, S2 - 6 hours and I3, S3 - 24 hours after the treatment. We assessed superoxid dismutase and glutathion peroxidase in the red blood cells. Both anesthetics drecreased the enzyme activities: isoflurane in groups I1 and I3, whereas sevoflurane in all experimental groups (S1, S2, S3).

Impact of Isoflurane and Sevoflurane Anesthesia on Kidney Structure and Function in Rats

Various researchers studied the nephrotoxicity of inhalational anesthetics, with contradictory results. We aimed to observe the impact of the repeated isoflurane or sevoflurane anesthesia on kidney structure and function. 40 female rats were used in this study. Animals were divided in 8 groups. Two groups served as control: IM (isoflurane) and SM (sevoflurane), exposed to oxygen alone (1l/min) 3 times, for 2 hours, every other day. Three groups were anesthetized with isoflurane (I1, I2, I3) and the other three (S1, S2, S3) with sevoflurane. The anesthetics were administered three times, for two hours long each, every other day. Rats were sacrificed through cervical dislocation as follows: groups IM, SM, I1, S1 imediately after the last anesthesia, groups I2, S2 – 6 hours post-anesthesia and groups I3, S3 – 24 hours post-anesthesia. Kidney samples were harvested, fixed in 10% buffered formaline, embedded in paraffin, sectioned (5 µm thickness) and stained with Goldner’s trichrome metod. After isoflurane anesthesia, the kidney displays a moderate congestion in the corticomedullary area, which maintains throughout the experimental period. The only changes in animals anesthetized with sevoflurane also appear on the vascular component. The congestion maintains throughout the experimental period in the renal cortex, as well as the corticomedullary area. No other aspects that suggest functional impairment are observed, in any of the experimental groups. Inhalational anesthesia with isoflurane and sevoflurane produces a moderate congestion, without majorly impairing the kidney function.

Evaluation of the amount of excess cement around the margins of cement-retained dental implant restorations: The effect of the cement application method

Complete removal of excess cement from subgingival margins after cementation of implant-supported restorations has been shown to be unpredictable. Remaining cement has been shown to be associated with periimplant inflammation and bleeding. The purpose of this study was to investigate and compare the amount of excess cement after cementation with 4 different methods of cement application for cement-retained implant-supported restorations. Ten implant replicas/abutments (3i) were embedded in acrylic resin blocks. Forty complete veneer crowns (CVCs) were fabricated by waxing onto the corresponding plastic waxing sleeves. The wax patterns were cast and the crowns were cemented to the implant replicas with either an interim (Temp Bond) or a definitive luting agent (FujiCEM). Four methods of cement application were used for cementation: Group IM-Cement applied on the internal marginal area of the crown only; Group AH-Cement applied on the apical half of the axial walls of the crown; Group AA-Cement applied to all axial walls of the interior surface of the crown, excluding the occlusal surface; and Group PI-Crown filled with cement then seated on a putty index formed to the internal configuration of the restoration (cementation device) (n=10). Cement on the external surfaces was removed before seating the restoration. Cement layers were applied on each crown, after which the crown was seated under constant load (80 N) for 10 minutes. The excess cement from each specimen was collected and measured. One operator performed all the procedures. Results for the groups were compared, with 1 and 2-way ANOVA and the Tukey multiple range test (α=.05). No significant difference in the amount of excess/used cement was observed between the 2 different types of cements (P=.1). Group PI showed the least amount of excess cement in comparison to other test groups (P=.031). No significant difference was found in the amount of excess cement among groups MI, AH, and AA. Group AA showed the highest amount of excess cement. The volume of cement used for group PI specimens was significantly higher than for those in the other groups (P=.001). With respect to the volume of cement loaded into the test crowns no statistically significant difference was observed among other test groups (groups IM, AH, and AA). Group MI used the least amount of cement, followed by group AH and AA. No correlation between the amount of used cement and the amount of excess cement was found in any of the tested groups. Within the limitations of this in vitro study, the least amount of excess cement was present when a cementation device was used to displace the excess cement before seating the crown on the abutment (Group PI). With this technique a uniform layer of the luting agent is distributed over the internal surface of the crown leaving minimal excess cement when the restoration is seated.

Hypoxia-Inducible Factor 1-Alpha Release After Intracoronary Versus Intramyocardial Stem Cell Therapy in Myocardial Infarction

We have investigated the effect of stem cell delivery on the release of hypoxia-inducible factor 1 alpha (HIF-1alpha) in peripheral circulation and myocardium in experimental myocardial ischemia. Closed-chest, reperfused myocardial infarction (MI) was created in domestic pigs. Porcine mesenchymal stem cells (MSCs) were cultured and delivered (9.8 +/- 1.2 x 10(6)) either percutaneously NOGA-guided transendocardially (Group IM) or intracoronary (Group IC) 22 +/- 4 days post-MI. Pigs without MSC delivery served as sham control (Group S). Plasma HIF-1alpha was measured at baseline, immediately post- and at follow-up (FUP; 2 h or 24 h) post-MSC delivery by ELISA kit. Myocardial HIF-1alpha expression of infarcted, normal myocardium, or border zone was determined by Western blot. Plasma level of HIF-1alpha increased immediately post-MI (from 278 +/- 127 to 631 +/- 375 pg/ml, p < 0.05). Cardiac delivery of MSCs elevated the plasma levels of HIF-1alpha significantly (p < 0.05) in groups IC and IM immediately post-MSC delivery, and returned to baseline level at FUP, without difference between the groups IC and IM. The myocardial tissue HIF-1alpha expression in the infarcted area was higher in Group IM than in Group IC or S (1,963 +/- 586 vs. 1,307 +/- 392 vs. 271 +/- 110 activity per square millimeter, respectively, p < 0.05), while the border zone contained similarly lower level of HIF-1alpha, but still significantly higher as compared with Group S. Trend towards increase in myocardial expression of HIF-1alpha was measured in Group IM at 24 h, in contrast to Group IC. In conclusion, both stem cell delivery modes increase the systemic and myocardial level of HIF-1alpha. Intramyocardial delivery of MSC seems to trigger the release of angiogenic HIF-1alpha more effectively than does intracoronary delivery.

Efficacy of a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine in pigs naturally exposed to a heterologous European (Italian cluster) field strain: Clinical protection and cell-mediated immunity

The purpose of this study was to assess clinical protection in pigs vaccinated with a commercially available attenuated porcine reproductive and respiratory syndrome virus (PRRSV) vaccine (Porcilis ® PRRS) and then naturally exposed under field conditions to a heterologous (Italian cluster) strain of virulent PRRSV. A total of 30, 4-week-old pigs seronegative for PRRSV were allocated to 1 of 3 groups (IM, ID, and C groups). At 5 weeks of age, pigs of groups IM ( n = 10 pigs) and ID ( n = 10 pigs) were vaccinated intramuscularly and intradermally, respectively, with modified live PRRSV-1 vaccine (Porcilis ® PRRS). Pigs of group C ( n = 10 pigs) were kept as non-vaccinated controls. At post-vaccination (PV) days 0, 7, 14, 28, and 45, blood samples were collected for detection of vaccine virus (PCR) and antibody response (ELISA), identification of changes in lymphocyte subpopulations by cytometry, and IFN-γ PRRSV-specific secreting cells (SC) by ELISpot. At PV day 45, pigs of A, B, and C groups were moved to a site 3 conventional finishing herd with a history of respiratory disease caused by PRRSV and the most common bacteria to be exposed to a natural challenge. The PRRSV field strain, belonging to the Italian cluster of the PRRSV-1, demonstrated a 84% identity with the vaccine virus (DV strain) at ORF5 sequencing. At 0 (exposure day = 45 days PV), 4, 7, 11, 14, 19, 21, 28, and 34 days post-exposure (PE) blood samples were collected for detection and titration of PRRSV and antibody, as well as for lymphocyte and IFN-γ measurement as described above. Throughout the post-exposure period, all pigs were observed daily for clinical signs. The overall clinical signs were reduced by 68 and 72%, respectively in the intramuscularly and intradermally vaccinated pigs compared to controls. Respiratory signs were reduced by 72 and 80%, respectively in the IM and ID groups. Clinical protection was associated with marked activation of cell-mediated immune response. The highest levels of specific IFN-γ production at 21–34 days PE were concomitant and associated to changes in natural killer (NK) cells, γ/δ T, and cytotoxic T lymphocytes in the blood. In our field study, evidences of EU attenuated vaccine-induced clinical protection against natural exposure to a genetically diverse (84% homology) PRRSV-1 isolate (Italian cluster) was demonstrated by the statistically significant reduction in clinical signs in terms of incidence, duration and severity and by a more efficient cell-mediated immune response in the vaccinated pigs as compared to the unvaccinated controls.

Effect of Eradication of Helicobacter pylori on the Histology and Cellular Phenotype of Gastric Intestinal Metaplasia

Eradication of Helicobacter pylori appears to reduce gastric cancer incidence. We examined the effect of successful H pylori therapy on histology, phenotype of gastric intestinal metaplasia (GIM) (complete vs incomplete), and expression of several biomarkers related to carcinogenesis. Ninety-six H pylori-positive patients from Japan were treated successfully and followed up prospectively over 4 years with yearly endoscopy and were classified into 3 groups: group CG, chronic gastritis without GIM (n = 36); group IM, chronic gastritis with GIM (n = 33); group DYS, and GIM with dysplasia/cancer in a different location of the stomach (n = 27). A total of 288 endoscopic procedures were performed. Histology, mucin-histochemistry, and immunoperoxidase assays using monoclonal antibodies (mAbs) for cell phenotype (monoclonal antibody Das-1/colonic) and for neoplasia (TC22 and p53) were performed. The GIM histologic score was higher in group DYS than in group IM (P < .05) and group CG (P < .0001). The GIM scores did not change in groups IM and DYS over 4 years. mAb Das-1 reactivity was higher in group DYS (63%) than in group IM (39%) and group GC (0%). After eradication of H pylori, mAb Das-1 reactivity disappeared in 40% of patients (P < .0001) despite the unchanged GIM scores, and regression of TC22-4 was noted in the same patients. H pylori eradication does not reduce the histologic GIM score, but changes the cellular phenotype of GIM. This change of phenotype may be an important factor in the reduction of cancer incidence after eradication of H pylori.

Effects of octreotide on epidermal growth factor receptor, tissue plasminogen activator, and plasminogen activator inhibitor during intraperitoneal adhesion formation.

Intraperitoneal adhesions remain a problem after abdominal surgery. Octreotide has been proved to be able to reduce the number, strength, and extent of fibrous bands at and away from the anastomotic site in an animal model of rats with intestinal resection and reanastomosis. The aim of the present study was to investigate whether epidermal growth factor receptor (EGF-R), tissue plasminogen activator (tPA), and plasminogen activator inhibitor (PAI) are involved in this process. Laparotomy with intestinal resection and reanastomosis was performed on 60 male Wistar rats. All rats were randomly assigned to five groups: receiving no medication (control; C), normal saline (NS), octreotide solution peritoneal irrigation (Oc), octreotide intramuscular injection (IM), and Oc plus octreotide intramuscular injection (Oc + IM). The concentrations of serum EGF-R, plasma tPA, PAI-1, and PAI-2, and the strength of wound healing were measured. The serum EGF-R concentration showed no significant change from the preoperative level in the C and NS groups 7 and 14 days after the abdominal surgery. However, it decreased significantly on postoperative days 7 and 14 in groups Oc, IM, and Oc + IM ( P < 0.05). The plasma tPA concentrations were significantly higher than the preoperative level in all groups of rats on postoperative day 7. The levels were higher in groups Oc, IM, and Oc + IM than in group C or group NS at that time ( P < 0.05). On postoperative day 14, the plasma tPA concentrations had returned to the preoperative level in group C and group NS. However, the concentrations in groups Oc, IM, and Oc + IM still remained at a significantly higher level than the concentrations in group C and group NS. The plasma PAI-1 and PAI-2 concentrations showed no significant difference from the preoperative level in group C and group NS on days 7 and 14 after the abdominal surgery. However, the concentrations in groups Oc, IM, and Oc + IM on postoperative days 7 and 14 were markedly lower than those in groups C and NS ( P < 0.05). The wound strength was significantly greater on day 14 than on day 7 in all groups. In the rats with octreotide irrigation, the EGF-R level was decreased, the plasma tPA concentration was higher, and the plasma PAI-1 and PAI-2 concentrations were lower when compared with values in group C and group NS rats on days 7 and 14 after surgery. The data suggest that EGF-R, tPA, PAI-1, and PAI-2 are all involved in the mechanism of octreotide's action in reducing adhesion formation.

Effect of Adrenocorticotropin on Milk and Plasma Cortisol and Prolactin Concentrations

Milk cortisol and prolactin concentrations were measured in 12 lactating Holstein cows for 3 days (Experiment 1). On day 3, cows were divided randomly into groups; IV1, IV4, IM and control. Group IV1 received one intravenous injections of 40 IU adrenocorticotropin, group IV4, the same dosage four times at 2-h intervals, group IM 240 IU adrenocorticotropin intramuscularly, and control cows were injected with saline. Cortisol concentrations in blood plasma were increased in all groups following adrenocorticotropin injection, while milk cortisol concentrations increased fourfold in groups IV4 and IM and remained unchanged in group IV1.In Experiment 2, infusion of 18% saline into the mammary gland of three cows increased the somatic cell count in the infused quarter but had no effect on prolactin and cortisol of milk or plasma.In Experiment 3, six cows each were assigned to treatment groups of saline controls, IV2–0 (40 IU adrenocorticotropin intravenously at 0 and 2h post-milking) or IV2–8 (40 IU adrenocorticotropin intravenously at 8 to 10h post-milking). Half udders of each cow were milked before treatment and 4h later. Increases of cortisol in plasma increased cortisol concentrations of milk 4h after each treatment (IV2–0 and IV2–8). However, by 12h after treatment in IV2–0 cortisol concentrations of milk had returned to normal.Increases in adrenal cortisol secretion are followed rapidly (within 4h) by increased cortisol concentrations in milk. However, these rapidly in the absence of sustanined increments in blood cortisol. Increased cortisol concentrations in milk most likely represent sustained elevations in plasma cortisol.