Sônia Maria da Silva

Ascorbic acid reduces gentamicin-induced nephrotoxicity in rats through the control of reactive oxygen species

BACKGROUND & AIM Oxidative stress has been implicated in the pathophysiology of many forms of acute renal failure. The aim was examine the effect of vitamin C on oxidative stress and its relationship with nitric oxide on gentamicin-induced nephrotoxicity in rats. METHODS We utilized 32 Wistar rats allocated in four groups of eight animals each: control (CTL), vitamin C (VIT C), gentamicin (GENTA), and GENTA + VIT C; all groups were treated during seven days. RESULTS Serum urea and creatinine, serum and renal tissue malondialdehyde, blood superoxide anion and hydrogen peroxide in GENTA were increased vs CTL and vs VIT C, and decreased in GENTA + VIT C vs GENTA (all P < 0.05). Serum nitric oxide increased in GENTA vs CTL and vs VIT C, and reduced in GENTA + VIT C vs GENTA (P < 0.001). Urinary nitric oxide was reduced in GENTA vs CTL and vs VIT C and increased in GENTA + VIT C vs GENTA (P < 0.001). Severe degeneration of proximal tubules was present in GENTA, but only mild lesions were observed in GENTA + VIT C. CONCLUSION This study suggests that VIT C is a valuable tool to protect against GENTA-induced nephrotoxicity, by reducing reactive oxygen species and increasing the nitric oxide.

Remote ischemic preconditioning on neovascularization and follicle viability on ovary autotransplantation in rats.

PURPOSE Verify the optimum remote vascular occlusion time to reduce ovarian injury in autologous transplants in rats. METHODS Twenty-four adult female rats were assigned to four groups: GC (control group): bilateral oophorectomy followed by ovary transplant; GIPC (ischemic preconditioning group): remote ischemic preconditioning at the iliac artery for 5, 10, and 15 minutes (GIPC-5, GIPC-20, and GIPC-15) previous to bilateral oophorectomy and ovarian transplantation. The right ovary was fixed in the retroperitoneum. Euthanasia was performed 4 days after the surgical procedure. The follicles were counted and classified as developing versus atretic. The immunohistochemical assay identified vascular factor of endothelial growth (VEGF) in the ovarian stroma and assessed the proliferation capacity by means of the Ki-67 in the ovarian follicles. RESULTS Every group showed an inflammatory infiltrate, luteous body, and ovarian follicles in several phases of development. The ischemic preconditioning groups displayed greater amounts of viable ovarian follicles and increased vascularization and vasodilatation than the control group. GIPC-15 showed the highest amount of viable follicles compared to the others (P < .05 GIPC-15 vs GC; GIPC-15 vs GIPC-5). More VEGF-labeled cells were observed in GIPC-10 than the control group (P < .05, GIPC-10 vs GC). The proliferation index assessed by Ki-67 marking showed GC: 80%; GIPC-5: 76%; GIPC-10: 67%; and GIPC-15: 64% (P > .05). CONCLUSIONS The PCI-15 cohort seem to be the most adequate timing to achieve functional support and preservation of a greater number of viable ovarian follicles.

Effect of Remote Ischemic Preconditioning on Rat Estradiol Serum Levels and Follicular Development After Ovarian Transplantation

PURPOSE The purpose of this study was to evaluate estradiol serum levels and follicular development in rats subjected to ovarian autologous transplantation with or without remote ischemic preconditioning (R-IPC). METHODS Seventy-two adult female Wistar EPM-1 rats were distributed into 3 groups: (1) controls; (2) ovarian transplantation; and (3) ovarian transplantation + R-IPC. The groups were divided into subgroups, according to the prefixed date for euthanasia: 24 hours, 48 hours, 72 hours, and 7th postoperative day (PO). R-IPC was performed by clamping the common iliac artery for a 15-minute period of ischemia followed by 15 minutes of reperfusion, before undergoing ovarian transplantation. The graft was fixed to the retroperitoneum with a simple 8-0 prolene thread. Blood samples were collected from the vena cava. For evaluation of follicular development, the ovarian follicles were classified as immature and mature follicles besides corpora lutea. Only the viable follicles and functioning corpora lutea were counted. RESULTS At 72 hours, the R-IPC group showed higher estradiol values than the other groups, which were similar. After 24 hours the mean values were similar among all groups, and at 48 hours the R-IPC group was similar to the transplanted group without IPC. Animals undergoing R-IPC showed superior morphologic aspects, but 7 days after transplantation the morphology was worse in all groups. R-IPC enhanced the number of immature follicles at 48 hours (P > .05) and number of mature follicles from 24 hours to 48 hours after transplantation (P < .01). Functioning corpora lutea number was increased as well. CONCLUSION R-IPC increased the estradiol levels in autologous ovarian transplants associated with better graft morphology and more mature follicles.

Study on the vaginal smear of rats submitted to autologous ovarian transplant: impact of remote ischemic preconditioning

PURPOSE To evaluate the remote ischemic preconditioning (R-IPC) impact on the quality of the ovarian graft by means of vaginal smear of transplanted rats. METHODS Sixty rats were used divided in six groups: Control; Fresh transplant (TxF); Cryopreserved transplant (TxC); R-IPC; R-IPC + fresh transplant (TxF+R-IPC); R-IPC + cryopreserved transplant (TxC+R-IPC). R-IPC was performed in the common iliac artery. Autologous ovarian tissue was implanted integrally in the retro peritoneum. On the first PO day, vaginal smear collection was daily initiated. After 30 days, the PO day when the estrous cycle was re-initiated was considered for analysis as well as the estrous days and the number of estrous cycles. RESULTS R-IPC showed a tendency to an early estrus re-initiation (p>0.05) as well as increase the number of cycles in the fresh transplanted group while in the cryopreserved transplant the number of cycles was similar, regardless of the stimulus R-IPC (p>0.05). The animals which had undergone fresh grafts had a longer estrous period than the ones which had undergone cryopreserved grafts, with or without R-IPC (p<0.05). CONCLUSION R-IPC promoted earlier re-initiation of ovarian activity in the PO and greater estrous frequency, with more consistent results in the fresh grafts than in the cryopreserved ones.

Effect of N-acetylcysteine in liver ischemia-reperfusion injury after 30% hepatectomy in mice

PURPOSE Evaluate the effect of N-acetylcysteine in liver remnant after hepatectomy associated to ischemia-reperfusion injury in mice. METHODS Male adult BALB/c mice, weighing 20-22 g were used. Animals were anesthetized with ketamine (70 mg/kg) and xylazine (10 mg/kg); received N-acetylcysteine (150 mg/kg, H-IR-NAC group) or vehicle (H-IR group). Surgical procedures were performed under 10X magnification. Partial hepatectomy (30%) was followed by ischemia-reperfusion injury (30 minutes of ischemia and 60 minutes of reperfusion). Blood sample and liver tissue were removed before animal was euthanized. AST and ALT were evaluated in blood samples and histomorphological analyses were performed in remnant liver. Groups were compared by Mann-Whitney test, and it was considered significant when p<0.05. RESULTS Biochemical evaluations showed reduced levels of ALT in NAC group (H-IR-NAC=376 ± 127 U/l vs H-IR=636 ± 39 U/l, p=0.023). AST was similar (p=0.456). H-IR group showed hepatic tissue with preserved architecture, large area of steatosis, vascular congestion and rare mitogenic activity. NAC group showed hepatic tissue with small area of steatosis, vascular congestion and elevated mitogenic activity, evidenced by increased binuclear cells (H-IR-NAC=15.88 ± 0.52 vs H-IR=7.4 ± 0.37, p<0.001). CONCLUSION N-acetylcysteine promotes enzymatic and morphological protection against hepatectomy and ischemia-reperfusion injury.

Progressive Evaluation of Apoptosis, Proliferation, and Angiogenesis in Fresh Rat Ovarian Autografts Under Remote Ischemic Preconditioning

This study evaluated the remote ischemic preconditioning (R-IPC) early and late repercussion on fresh ovarian transplants, aiming to assess a probable protective effect in ovarian follicular pool. Sixty Wistar EPM-1 rats were used, divided in 2 study groups: ovarian transplantation (Tx) and Tx + R-IPC, submitted to ovary transplant with or without R-IPC, respectively. These groups were subdivided according to the date for euthanasia: 4th, 7th, 14th, 21st, and 30th days of the postoperatory period. Morphology, morphometry, neoangiogenesis (vascular endothelial growth factor [VEGF]), proliferative activity (Ki-67), and apoptosis (cleaved caspase-3) were evaluated. Remote ischemic preconditioning was performed in the common iliac artery. Fresh autologous ovarian tissue was implanted integrally in the retroperitoneum. All animals showed resumption of estrous phase after ovary transplantation. Remote ischemic preconditioning attenuated the lesions progressively from the 7th day, with greater number of the immature follicles (14 days, P < .05), but didn’t affect mature follicles and corpora lutea (P > .05). Immunohistochemical analyzes, taken as a whole, show that R-IPC benefic effect is more evident in the later periods of evaluation, when a greater proliferative activity (14, 21, and 30 days, P < .05) and lesser cell apoptotic activity (21 and 30 days, P < .05). The VEGF expression was similar in all times (P > .05). Remote ischemic preconditioning could have a benefic effect in the progressive evaluation of freshly grafted ovarian, especially on the latest phases of the posttransplant period. The 14th day was a landmark in the recuperation of the graft. Further investigations are necessary to determine the role of R-IPC in this scenario and its effect in frozen-thawed ovarian tissue.

Effect of N-acetylcysteine in hearts of rats submitted to controlled hemorrhagic shock

Introduction Pharmacological therapy is a strategy for the prevention of complications associated with ischemia and reperfusion injury that occurs after volume replacement in the treatment of hemorrhagic shock. Objective The aim of this study was to evaluate the effect of N-acetylcysteine associated with fluid resuscitation in cardiac injury in a rat hemorrhagic shock model. Methods Mice Wister male rats were randomly and subjected to controlled hemorrhagic shock for 60 min. and then, subjected to resuscitation with Ringer lactate. In a group of six animals, 150mg/kg of N-acetylcysteine were added to fluid volume replacement. The animals were observed for 120 min and after this period, were euthanized and cardiac tissue was collected for histopathological analysis and measurement of thiobarbituric acid reactive substances and pro-and anti-inflammatory interleukin. Results Cardiac tissue of the group treated with N-acetylcysteine showed lower concentrations of thiobarbituric acid reactive substances (0.20±0.05 vs. 0.27±0.05, P=0.014) and reduced histopathological damage and edema when compared to the group whose volume replacement occurred only with Ringer lactate. There was no difference in the expression of cytokines interleukin 6 (2,138.29±316.89 vs. 1,870.16±303.68, P=0.091) and interleukin 10 (1.019,83±262,50 vs. 848.60±106.5, P=0.169) between the treated groups. Conclusion The association of N-acetylcysteine on volume replacement attenuates oxidative stress in the heart, as well myocardial damage and edema, but does not modify the expression of inflammatory cytokines.

Indirect evaluation of estrogenic activity post heterotopic ovarian autograft in rats

PURPOSE To morphologically evaluate the estrogenic effect on the uterus and vagina of rats submitted to ovarian autografts. METHODS Twenty Wistar EPM-1 adult rats were bilaterally ovariectomized, followed by ovarian transplants in retroperitoneal regions. The animals were divided in four groups of five animals, according to the day of euthanasia: G4, G7, G14 and G21, corresponding to the 4th, 7th, 14th and 21st day after surgery, respectively. Vaginal smears were collected from the first day of surgery until euthanasia day. After that, the vagina and uterus were removed, fixed in 10% formaldehyde and submitted to histological analysis and stained with hematoxiline and eosine. RESULTS All animals showed estrous cycle changes during the experiment. In 4th day, the uterus showed low action of estrogen with small number of mitosis and eosinophils as well as poor development. On the 7th day, the endometrium was atrophic without mitotic signals and presented a small number of eosinophils. On the 14th and 21th days the histological findings were similar, with the presence of mitosis in the endometrial glands and intense leucocyte infiltration with a large number of eosinophils. Morphometric results showed that the endometrial and myometrial thickness as well as the number of eosinophils presented the highest values during the 14th and 21th days of the evaluation. The 7th day group also presented the lowest eosinophil numbers. Vaginal epithelium features were: 4th and 7th day groups presented non-keratinized stratified epithelium with 5 and 2 cell layers, respectively. The 14th and 21st day groups presented non-keratinized stratified epithelium with 14 and 15 cell layers. CONCLUSION Experimental ovarian autografts in the evaluated organs presented maximum estrogen activity after the 21st day of surgery, according to morphological and morphometric data.

Effect of N-acetylcysteine in rat hearts subjected to controlled hemorrhagic shock

Introduction Pharmacological therapy is a strategy for the prevention of complications associated with ischemia and reperfusion injury that occurs after volume replacement in the treatment of hemorrhagic shock. Objective The aim of this study was to evaluate the effect of N-acetylcysteine associated with fluid resuscitation in cardiac injury in a rat hemorrhagic shock model. Methods Mice Wister male rats were randomly and subjected to controlled hemorrhagic shock for 60 min. and then, subjected to resuscitation with Ringer lactate. In a group of six animals, 150mg/kg of N-acetylcysteine were added to fluid volume replacement. The animals were observed for 120 min and after this period, were euthanized and cardiac tissue was collected for histopathological analysis and measurement of thiobarbituric acid reactive substances and pro-and anti-inflammatory interleukin. Results Cardiac tissue of the group treated with N-acetylcysteine showed lower concentrations of thiobarbituric acid reactive substances (0.20±0.05 vs. 0.27±0.05, P=0.014) and reduced histopathological damage and edema when compared to the group whose volume replacement occurred only with Ringer lactate. There was no difference in the expression of cytokines interleukin 6 (2,138.29±316.89 vs. 1,870.16±303.68, P=0.091) and interleukin 10 (1.019,83±262,50 vs. 848.60±106.5, P=0.169) between the treated groups. Conclusion The association of N-acetylcysteine on volume replacement attenuates oxidative stress in the heart, as well myocardial damage and edema, but does not modify the expression of inflammatory cytokines.