Effects of rivaroxaban on myocardial mitophagy in the rat heart.

Background: This study aims to demonstrate the efficacy of rivaroxaban's pharmacokinetic effects on myocardial mitophagy in rats by inducing apoptosis. Methods: In this double-blind experiment, Wistar albino male rats were randomly divided into three groups for an experimental ischemia model: the sham group (Group 1; n=7), the control group (Group 2; n=7), and the drug group (Group 3; n=7). Rivaroxaban was perorally administered with gavage at 2 mg/ kg/day for 28 days in Group 3. The heart was surgically exposed, and ischemia was achieved by compressing the vessel around the proximal part of the left anterior descending coronary artery for 10 min. The heart tissue was then transected, removed, and morphologically and immunohistochemically examined under a light microscope. Results: Heart sections were immunohistochemically marked with caspase 3, caspase 9, APAF1, and Bcl-2 antibodies. Group 1 was compared to the rivaroxaban-treated group, and the pathways inducing apoptosis was increased (caspase 3, caspase 9, APAF1; p<0.015, p<0.004, and p<0.01, respectively) and Bcl-2, the molecule that inhibits apoptosis, was decreased (p<0.01) in Group 3. Conclusion: The present study provides an evidence that the mitophagy response is less in rivaroxaban-treated rats, showing the protective effect of rivaroxaban against acute ischemia. Rivaroxaban-treated rats may have reduced cell death in cardiomyocytes during myocardial infarction and thus have reduced damage to the heart tissue caused by myocardial infarction.

Histomorphological Investigation of Microfracture Location in a Rabbit Osteochondral Defect Model.

BACKGROUND: Microfracture is the most common treatment for cartilage defects of the knee. In microfracture surgery, holes are randomly drilled into the subchondral bone. The effect of the hole's location on its interaction with the cartilage defect site and its influence on the healing process is currently uncertain. PURPOSE: To investigate the effects of different microfracture locations on healing in a rabbit knee osteochondral defect model. STUDY DESIGN: Controlled laboratory study. METHODS: A total of 29 adult New Zealand White rabbits were divided into 5 groups. In the healthy cartilage control group (n = 5), no surgical procedure was performed. Cylindrical full-thickness cartilage defects (5 × 3 mm) were created in the patellar groove of the remaining 24 rabbits. In the defect control group (n = 6), only the defect was created. A microfracture was performed at the 12-o'clock position (group peripheral single; n = 6), centrally (group central; n = 6), and at the 12- and 6-o'clock positions (group peripheral double; n = 6) of the defect. The animals were sacrificed after 8 weeks. Cartilage healing was evaluated by International Cartilage Regeneration & Joint Preservation Society (ICRS) score, modified O'Driscoll score, immunohistochemical analysis (type 1 collagen, type 2 collagen, and aggrecan), and scanning electron microscopy analysis. RESULTS: In group peripheral double, better cartilage healing was observed in all parameters compared with the other groups (P < .05). Group peripheral double had the greatest amount of filling, with 79% of the defect area filled with fibrocartilage repair tissue. Group peripheral single demonstrated filling of 73% of the defect area, group central 56%, and the defect control group 45%. The ICRS score was significantly higher in group peripheral single compared with group central and the defect control group. Type 2 collagen and aggrecan immunoreactivity were significantly stronger in group central than group peripheral single and the defect control group (P < .05). CONCLUSION: Microfracture performed at the peripheral margin of the defect had better filling characteristics in a rabbit model. This study suggests that interaction of pluripotent cells released from the microfracture site with the intact cartilage may enhance the quality of the repair tissue. CLINICAL RELEVANCE: The location of microfracture holes in relation to the peripheral border of the osteochondral defect (to the intact cartilage) is important in both the quality and the quantity of the newly formed repair tissue.

The healing effect of topical tea tree oil on pressure ulcers in a rat model.

Objective: The effects of topical tea tree oil (TTO) on the healing of pressure ulcers (PUs) in an animal model was evaluated. Method: To induce PUs, ischaemia-reperfusion cycles were performed by the external application of magnetic plates, with an ischaemic period of eight hours and a reperfusion period of 16 hours. Male and female Wistar rats were divided into three equally sized groups (n=20): one group received topical glycerin twice daily, another group received topical 10% (volume/volume (v/v)) TTO in glycerin twice daily; and the remaining group was untreated. The animals were assessed after one, four, seven and 14 cycles of ischaemia-reperfusion by thermal camera imaging, and then euthanised and sampled to investigate the degree of inflammation, collagen synthesis and apoptosis in the PUs. Results: Although topical glycerin alone suppressed local inflammation and apoptosis, this suppressive effect was accentuated at all timepoints by the application of topical TTO + glycerin. Similarly, an increase in collagen synthesis was observed in the glycerin group and this was accentuated by TTO at all timepoints. Parallel to the histological findings, the local temperature had decreased significantly on days 4 and 7 for both treatment groups (glycerin and TTO+glycerin). Conclusion: In this study, treatment with 10% (v/v) TTO in glycerin effectively suppressed skin inflammation and apoptosis, while it increased collagen synthesis during PU formation.

Loofah-chitosan and poly (-3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) based hydrogel scaffolds for meniscus tissue engineering applications.

The meniscus is a fibrocartilaginous tissue that is very important for the stability of the knee joint. However, it has a low ability to heal itself, so damage to it will always lead to articular cartilage degeneration. The goal of this study was to make a new type of meniscus scaffold made of chitosan, loofah mat, and PHBV nanofibers, as well as to describe hydrogel composite scaffolds in terms of their shape, chemical composition, mechanical properties, and temperature. Three different concentrations of genipin (0.1, 0.3, and 0.5 %) were used and the optimal crosslinker concentration was 0.3 % for Chitosan/loofah (CL) and Chitosan/loofah/PHBV fiber (CLF). Scaffolds were seeded using undifferentiated MSCs and incubated for 21 days to investigate the chondrogenic potential of hydrogel scaffolds. Cell proliferation analyses were performed using WST-1 assay, GAG content was analyzed, SEM and fluorescence imaging observed morphologies and cell attachment, and histological and immunohistochemical studies were performed. The in vitro analysis showed no cytotoxic effect and enabled cells to attach, proliferate, and migrate inside the scaffold. In conclusion, the hydrogel composite scaffold is a promising material for engineering meniscus tissue.

Fabrication of 3D Printed poly(lactic acid) strut and wet-electrospun cellulose nano fiber reinforced chitosan-collagen hydrogel composite scaffolds for meniscus tissue engineering.

The main goal of the study was to produce chitosan-collagen hydrogel composite scaffolds consisting of 3D printed poly(lactic acid) (PLA) strut and nanofibrous cellulose for meniscus cartilage tissue engineering. For this purpose, first PLA strut containing microchannels was incorporated into cellulose nanofibers and then they were embedded into chitosan-collagen matrix to obtain micro- and nano-sized topographical features for better cellular activities as well as mechanical properties. All the hydrogel composite scaffolds produced by using three different concentrations of genipin (0.1, 0.3, and 0.5%) had an interconnected microporous structure with a swelling ratio of about 400% and water content values between 77 and 83% which is similar to native cartilage extracellular matrix. The compressive strength of all the hydrogel composite scaffolds was found to be similar (∼32 kPa) and suitable for cartilage tissue engineering applications. Besides, the hydrogel composite scaffold comprising 0.3% (w/v) genipin had the highest tan δ value (0.044) at a frequency of 1 Hz which is around the walking frequency of a person. According to the in vitro analysis, this hydrogel composite scaffold did not show any cytotoxic effect on the rabbit mesenchymal stem cells and enabled cells to attach, proliferate and also migrate through the inner area of the scaffold. In conclusion, the produced hydrogel composite scaffold holds great promise for meniscus tissue engineering.

Angiotensin receptor-neprilysin inhibition by sacubitril/valsartan attenuates doxorubicin-induced cardiotoxicity in a pretreatment mice model by interfering with oxidative stress, inflammation, and Caspase 3 apoptotic pathway.

OBJECTIVE: Doxorubicin (DOX) is a well-known cardiotoxic agent, whereas sacubitril/valsartan (Sac/Val) is an effective treatment option in heart failure. In this study, we aimed to evaluate the effect of Sac/Val on DOX-induced cardiotoxicity in pretreatment mice model. METHODS: A total of 24 mice were equally classified into 4 groups; control group, DOX (20 mg/kg; fifth day), Sac/Val (80 mg/kg), and Sac/Val+DOX (Sac/Val was given from day one of the study before doxorubicin administration). Electrocardiography parameters, including durations of QRS, ST, QT, PP segment, and QT/PQ index were measured. Total antioxidant status (TAS), total oxidant status (TOS), tumor necrosis factor-α (TNF-α), interleukin 1ß (IL-1ß), IL-6, NT-proBNP concentrations, and Caspase 3 activity were evaluated. RESULTS: At the end of the 9-day study duration, QRS, ST, QT intervals, QT/PQ index and TAS, TOS, TNF-α, IL-1ß, IL-6 levels were significantly higher in the DOX group than in the control group (p<0.001). Moreover, there were significant differences only in the PP interval when comparing the Sac/Val+DOX and control groups (p<0.001). QRS, ST, QT intervals, and QT/PQ index, TAS, TOS, TNF-α, IL-1ß, IL-6 levels were significantly lower in the Sac/Val+ DOX group compared with the DOX group (p<0.001). Furthermore, NT-proBNP levels were lower in the Sac/Val+DOX group compared with the DOX group along with less Caspase 3 apoptosis. CONCLUSION: Sac/Val seems to be cardioprotective against DOX-induced cardiotoxicity in pretreatment mice model. These findings can be attributed to the antiarrhythmic, anti-inflammatory, antioxidant, and antiapoptotic effects of Sac/Val as shown in this study.

Effect of bivalirudin on neointimal hyperplasia and endothelial proliferation in rabbit.

BACKGROUND: The aim of this study was to investigate the effects of bivalirudin on endothelial cell proliferation and neointimal hyperplasia in a rabbit carotid artery model. METHODS: "New Zealand rabbits (n = 12)" weighing 2-3 kg were randomly divided into two groups. Arteriotomy was performed to the rabbit carotid artery and closed with continuous suture technique. Group B (n = 6) as a control group received 150 U/kg heparin sodium; however, group A (n = 6) was given 0.75 mg/kg bivalirudin i.v. bolus and infusion 1.75 mg/kg/hour (B01AE06-Bivalirudin 250 mg) during perioperation period. At the end of the 28th day, the carotid artery segment was excised and evaluated histologically. RESULTS: All histological and immune staining analyzes were performed by two blind researchers in the treatment of rabbits. In the control group rabbit carotid artery sections, tunica intima was observed to thicken. In the bivalirudin group, intimal hyperplasia was less observed compared to the control group. No significant difference was observed between groups in tunica media thickness. Lumen diameter and lumen area were found to be wider in the experimental group. P value was found to be less than 0.05. CONCLUSION: Our study demonstrates that bivalirudin significantly affects and prevents neointimal hyperplasia and endothelial cell proliferation.

Epigenetic Programming Through Breast Milk and Its Impact on Milk-Siblings Mating.

BACKGROUND: The epigenetic effects of transmission of certain regulatory molecules, such as miRNAs, through maternal milk on future generations, are still unknown and have not been fully understood yet. We hypothesized that breastfeeding regularly by adoptive-mother may cause transmission of miRNAs as epigenetic regulating factors to the infant, and the marriage of milk-siblings may cause various pathologies in the future generations. RESULTS: A cross-fostering model using a/a and A vy /a mice had been established. F2 milk-sibling and F2 control groups were obtained from mating of milk-siblings or unrelated mice. Randomized selected animals in the both F2 groups were sacrificed for miRNA expression studies and the remainings were followed for phenotypic changes (coat color, obesity, hyperglycemia, liver pathology, and life span). The lifespan in the F2 milk-sibling group was shorter than the control group (387 vs 590 days, p = 0.011) and they were more obese during the aging period. Histopathological examination of liver tissues revealed abnormal findings in F2 milk-sibling group. In order to understand the epigenetic mechanisms leading to these phenotypic changes, we analyzed miRNA expression differences between offspring of milk-sibling and control matings and focused on the signaling pathways regulating lifespan and metabolism. Bioinformatic analysis demonstrated that differentially expressed miRNAs were associated with pathways regulating metabolism, survival, and cancer development such as the PI3K-Akt, ErbB, mTOR, and MAPK, insulin signaling pathways. We further analyzed the expression patterns of miR-186-5p, miR-141-3p, miR-345-5p, and miR-34c-5p and their candidate target genes Mapk8, Gsk3b, and Ppargc1a in ovarian and liver tissues. CONCLUSION: Our findings support for the first time that the factors modifying the epigenetic mechanisms may be transmitted by breast milk and these epigenetic interactions may be transferred transgenerationally. Results also suggested hereditary epigenetic effects of cross-fostering on future generations and the impact of mother-infant dyad on epigenetic programming.

Efficiency of Minimized Circuits of a Heart Roller Pump on Systemic Inflammatory Response Syndrome and Multiorgan Effects in a Rat Model.

BACKGROUND: The aim of this study is to compare the effects of tubing length on systemic inflammatory response syndrome and myocardial protection in a rat model of cardiopulmonary bypass (CPB) from a histological standpoint. METHODS: Twelve adult male Wistar Albino rats weighing >180 g were randomly selected and divided into 2 groups. In 1 group, the pump lines were kept 1 m shorter than standard. The right jugular vein and tail artery were cannulated using a 16-gauge catheter. Animals received 500 IU/kg intravenous heparin. Cardiac index and rectal temperature were set at 2.4 mL and 36°C, respectively. Total line volume was maintained at 8 mL. A roller pump was adjusted to supply a blood flow of 6 to 28 mL/min (mean 10 mL/min), similar to the typical cardiac output of rats. CPB duration was 15 minutes throughout the experiment. After sacrifice, tissue samples were collected from heart, liver, and kidney for histomorphologic examination. RESULTS: All histochemical and histomorphologic analyses, performed by 2 blinded researchers, revealed band loss in cardiomyocytes, mononuclear (MNL) cell infiltration, and impaired fibrillar organization in the standard-line group. Additionally in that group, sinusoidal dilatation in the liver, low-level congestion, focal necrosis, and periportal MNL infiltration were noted. In the shorter-line group, on the other hand, MNL cell infiltration, band loss in myofibrils, and cardiomyocyte degeneration were rarely observed. Higher liver congestion and lower MNL cell infiltration were observed in the shorter-line group. No significant differences were found in kidney samples. CONCLUSION: In a shorter-line roller pump test model, less multiorgan damage and fewer systemic inflammatory responses were observed. It may be applicable to keep CPB lines as close to the table as possible, especially in pediatric cardiac surgery cases.

Effects of carbon dioxide insufflation on anastomosis remodeling at a carotid artery site in rabbits.

INTRODUCTION: Use of carbon dioxide (CO2) insufflation (CDI) on the surgical field during heart surgery has become widespread, and in some units routine. AIM: To assess the effects of CDI on endothelial dysfunction in a carotid artery model in rabbits. MATERIAL AND METHODS: Twelve randomly selected rabbits were divided into two groups. Right carotid arteries of the animals were transected and sutured with running suture technique. Then, 1 l/min CO2 insufflation was initiated with a 45° angle. In the control group, the anastomotic field was irrigated with 0.1 ml/s flow of 0.9% saline. At day 28, the carotid artery segments were removed and prepared for histological specimens. RESULTS: In the cross-sections of the control group vessel samples, thickening of the tunica intima was observed. Scoring the quantity of endothelial nitric oxide synthase (e-NOS) and α-smooth muscle actin (α-SMA) positive staining revealed a nonsignificant difference between the experimental groups (p = 0.07). In the CO2 group, the intimal hyperplasia (p = 0.2) and the thickness of the tunica media (p = 0.2) were indistinguishable when compared to the control group. The mean luminal diameters and luminal areas of the experimental groups were all evaluated by histomorphometry and a significant differences was found between luminal areas (p = 0.016). On the other hand, no significant difference was found between mean luminal diameters (p = 0.055). CONCLUSIONS: Our study indicated that CDI can affect endothelial cell damage and the mean luminal diameters.

Effects of rivaroxaban on intimal hyperplasia and smooth muscle cell proliferation at the carotid artery anastomosis site in rabbits.

OBJECTIVE: This study aimed to assess the effect of new generation oral, direct factor Xa inhibitor rivaroxaban on intimal hyperplasia and smooth muscle cell proliferation at the carotid artery anastomosis site of rabbits. METHODS: In total, 14 New Zealand male rabbits weighing 3-3.5 kg were randomized into two groups. Group A (7 rabbits) served as the control group and received no medication. Rivaroxaban was perorally administered to group B (7 rabbits) mg/kg/day for 28 days. Following anesthesia induction, carotid arteries were dissected through a right neck incision. following heparinization at 100 IU/kg, vertical full thickness arteriotomy was performed, then was repaired continuously with 8-0 polypropylene. At day 28, all rabbits were sacrificed and the anastomosed carotid artery segments were analyzed using light microcopy. Hematoxylin-eosin and Masson's trichrome stained images were analyzed using a digital image analysis program, and lumen diameter, lumen area, intimal and medial thickness, and media areas were measured and results were compared. RESULTS: In the serial sections, the average lumen diameter of group B was higher than that of group A (p=0.001). The lumen areas of group B were also higher than those of group A (p=0.004). The intimal thickness of group B was lower than that of group A (p=0.001). When the section series were evaluated for media thickness, the thickness of group B was lesser than that of group A; the difference was statistically significant (p=0.002). CONCLUSION: This study may imply a potential midterm benefit of rivaroxaban following arterial anastomosis by reducing intimal proliferation and restenosis.

The effects of α-lipoic acid on aortic injury and hypertension in the rat remnant kidney (5/6 nephrectomy) model.

OBJECTIVE: The present study was designed to investigate the effects of α-lipoic acid on the abdominal aorta and hypertension in a remnant kidney model histomorphometrically, immunohistochemically, and ultrastructurally. METHODS: We surgically reduced the renal tissue mass to 5/6 by applying a remnant kidney model. The rats were divided into 4 groups: Group 1- control group, Group 2- lipoic acid group, Group 3- 5/6 nephrectomy group, and Group IV: 5/6 nephrectomy+lipoic acid-treated group. Lipoic acid solution 100 mg/kg was administered by oral gavage for 8 weeks to Groups II and IV. At the end of the experiment, systemic mean blood pressure was monitored. Then, aortic tissues were removed and fixed. After routine histological procedures, tissue sections were examined histochemically, immunohistochemically (type I angiotensin receptor, vascular endothelial growth factor, alpha-smooth muscle actin), and ultrastructurally. RESULTS: The blood pressure measurements in 5/6 nephrectomy group were significantly higher compared to other groups. In the 5/6 nephrectomy+lipoic acid group, measured blood pressure values and tunica media thickness were significantly lower than in the 5/6 nephrectomy group. In the 5/6 nephrectomy+lipoic acid group, decreased aortic wall thickness, regularity in the structure of elastic fibrils, and more organized elastic lamellae were seen. The expression of type I angiotensin receptor, vascular endothelial growth factor, alpha-smooth muscle actin in the 5/6 nephrectomy+lipoic acid group was decreased compared to the 5/6 nephrectomy group. CONCLUSION: In the present study, we found that α-lipoic acid could be a favorable agent for the target organ effects of secondary hypertension.

The effect of lipoic acid on wound healing in a full thickness uterine injury model in rats.

Aim of this study was to investigate the effects of lipoic acid on uterine wound healing by immunohistochemical and biochemical assay in a rat uterine horn model with full thickness injury. Thirty-two female Wistar albino rats were randomised into five groups: Control group, with no intervention; uterine scar group 15days (US15d), uterine scar group 15 days + alpha lipoic acid (ALA) (US15d + ALA), uterine scar group 30 days (US30d) and uterine scar group 30 days + ALA (US30 days + ALA). After uterine incision 100 mg/kg of ALA was administered by oral gavage for either 15 or 30 days. Vascular endothelial growth factor (VEGF) and alpha smooth muscle actin (α-SMA) distribution were evaluated by immunohistochemical methods in tissue and ELISA methods in tissue homogenate. The percentage of α-SMA positive area in US15d + ALA and US30d + ALA groups was significantly higher than US15 and US30d groups. The percentage of VEGF positive area in US15d + ALA group was significantly higher than US15d group and US30d + ALA group was significantly higher than US30d group. Biochemically, α-SMA was significantly higher in the US15d + ALA group when compared to US15d group and higher in US30d + ALA group when compared to US30d group. VEGF was significantly higher in US15d + ALA and US30d + ALA groups when compared to US15 and US30d groups. In conclusion, ALA was found to be effective in enhancing wound healing in uterine full thickness injury.

Lipoic acid decreases peritoneal adhesion formation in a rat uterine scar model.

OBJECTIVE: To investigate the effects of lipoic acid in the prevention of postoperative pelvic adhesions by a visual scoring system and immunohistochemistry in a rat uterine horn model with full thickness injury. MATERIAL AND METHODS: Twenty-eight female Wistar albino rats were randomised into four groups: uterine trauma control, 15 days and 30 days, and uterine trauma + lipoic acid, 15 days and 30 days. A full thickness defect was established by incising a segment of approximately 1.0 cm in length from each uterine horn, leaving the mesometrium intact. Extension and severity of the adhesions in each group were scored by a visual scoring system and evaluated immunohistochemically. RESULTS: Adhesion scores were 2.00±0.81, 2.14±0.69 0.71±0.75, and 0.85±0.69 for extent and 2.28±0.48, 2.14±0.69, 0.85±0.69, and 1.14±0.69 for severity in Groups 1, 2, 3 and 4, respectively. Adhesion extent and severity were significantly less for groups treated by lipoic acid but no difference was observed between long and short administration. Both Vitronectin and u-PAR staining were significantly increased in treatment groups when compared to the control group. CONCLUSION: Lipoic acid was found to be effective in reducing postoperative adhesion formation in a rat model.