Preoperative and postoperative administration of hyperbaric oxygen improves biochemical and mechanical parameters on ischemic and normal colonic anastomoses.

Colonic anastomotic leaks are a major postoperative complication, causing increased mortality and morbidity. Tissue ischemia is one of the most important factors that disrupt anastomotic healing. It is possible to reverse inadequate tissue oxygenation by using increased atmospheric pressure and hyperoxia, which are obtained from hyperbaric oxygen treatment (HBO). Our aim was to investigate the effects of preoperative and postoperative HBO treatment on normal and ischemic colonic anastomoses in rats. Eighty male Wistar Albino rats, weighing between 180 and 240 g, were divided into 8 equal groups. A 1-cm segment of left colon was resected 3 cm proximal to the peritoneal reflection in all groups and colonic anastomosis was performed. In groups 2, 4, 6 and 8, colonic ischemia was established by ligating 2 cm of mesocolon on either side of the anastomosis. Control groups (1 and 2) received no HBO. HBO treatment was given preoperatively in groups 3 and 4, postoperatively in groups 5 and 6, and both preoperatively and postoperatively in groups 7 and 8. HBO treatment was applied for 2 days in the preoperative period and 4 days in the postoperative period. Relaparotomy was performed on postoperative day 5 and a perianastomotic colon segment 2 cm in length was excised for detection of biochemical and mechanical parameters of anastomotic healing and histopathological evaluation. HBO treatment increased tissue hydroxyproline levels in all groups, and this difference was significant in normal anastomosis groups receiving preoperative HBO compared to controls (p = .013 for group 1 vs. group 3; p = .023 for group 1 vs. group 5). This improvement was more evident in ischemic and normal groups treated by administration of combined pre- and postoperative HBO (p = .021 and p = .013). HBO treatment also increased the mean bursting pressure values in all groups, and again, a statistically significant increase was noted in the ischemic groups compared to controls (p = .002 for group 2 vs. group 6; p = .001 for group 2 vs. group 8). Histopathological evaluation of anastomotic line fibrosis was not found to show significant differences between the groups. Adequate tissue oxygenation is the main factor in wound and anastomosis healing. HBO treatment has a positive effect on biochemical and mechanical parameters of ischemic and normal colon anastomoses in rats. It is possible to see this effect more clearly with combined HBO treatment applied before and after ischemic anastomosis.     

Sildenafil accelerates liver regeneration after partial hepatectomy in rats

Objective

The regeneration process causes the liver to achieve an adequate volume and function after major hepatectomy or living donor liver transplantation. Sildenafil, a selective phosphodiesterase-5 inhibitor used for erectile dysfunction, impacts the liver by enhancing the effects of nitric oxide. The aim of this study was to investigate the influence of sildenafil on liver regeneration in rats after partial hepatectomy.

Methods

Sixty young female Wistar Albino rats were randomly divided into three equal groups before 70% hepatectomy. Thereafter, we administered intraperitoneal saline to the control group (G1); 10 μg/kg sildenafil to the low-dose group (G2) and 100 μg/kg to the high-dose sildenafil group (G3). Half of the rats per group were sacrificed on the first and the other half on the fifth postoperative day after partial hepatectomy. Regeneration was assessed using three methods: (1) the formula described by Kwon et al formula, (2) the average number of mitotic figures in 10 microscopic fields, and (3) the average of Ki-67-positive nuclei in 1000 cells using immunohistochemistry.

Results

Although, the hepatic regeneration and mitosis rates were similar in all three groups, Ki-67 levels were significantly higher in both G2 and G3 than the control group on the first postoperative day. Hepatic regeneration was significantly greater in G2 and G3 than the control group as was the mitosis rate in the G2 group versus the two groups. By the 5th postoperative day Ki-67 levels were similar in the three groups.

Conclusion

Sildenafil treatment accelerated hepatic regeneration after partial hepatectomy in rats.     

Oral sildenafil citrate as an effective alternate in the treatment of postoperative pulmonary hypertensive crisis after congenital heart surgery

A five-month-old girl of Down syndrome underwent a corrective surgery for complete atrioventricular septal defect. Her postoperative course was complicated with pulmonary hypertensive (PH) crises despite nitroglycerin (NTG) infusion and inhaled nitric oxide (NO). Sildenafil citrate, a phosphodiesterase 5 inhibitor, was administered through a nasogastric tube at a starting dose of 0.3 mg/kg by stepwise increment to the maximum dose of 2 mg/kg 4 hourly. Sildenafil citrate dramatically lowered pulmonary arterial pressure and the patient was weaned from NTG and NO without PH crisis. There was no side effect after sildenafil citrate administration. Oral sildenafil citrate is a safe and potent adjunct to the existing therapies for postoperative PH in infants after open heart surgery.     

The effect of remote ischemic preconditioning on healing of colonic anastomoses

BACKGROUND: We aimed to investigate the potential protective effect of remote ischemic preconditioning (IPC) on delayed colonic anastomotic healing induced by remote ischemia and reperfusion (I/R) injury. MATERIALS AND METHODS: Forty male Wistar rats were randomly assigned into four groups, each consisting of 10 rats: the control group (C), the remote I/R group [I/R, 40 min of superior mesenteric artery (SMA) occlusion], the preconditioned I/R group (IPC, two cycles of 5 min temporary occlusion of SMA before an ischemic insult of 40 min), and the preconditioned group (PC, two cycles of 5 min temporary occlusion of SMA). Colonic anastomosis was performed immediately after the ischemic insult. Anastomotic healing was assessed on postoperative day 7 by determining anastomotic bursting pressure (ABP), tissue hydroxyproline content, histopathological examination, malondialdehyde (MDA), and nitric oxide levels. RESULTS: Remote I/R injury resulted with significant impairment in anastomotic healing in terms of mean ABP (P = 0.004), hydroxyproline content (P = 0.002), histopathological healing score (P = 0.001), nitric oxide level (P = 0.010), and MDA levels (P = 0.0001) when compared with the control group, but remote IPC did not improve all above mentioned parameters (P = NS for all), except MDA level (P = 0.011) when compared with I/R group. PC alone impaired the ABP (P = 0.0001), but it did not significantly change the other parameters measured (P = NS). CONCLUSIONS: The results of this study showed that remote IPC did not prevent I/R-induced delaying in colonic anastomotic healing.     

Locally applied molgramostim improves wound healing at colonic anastomoses in rats after ligation of the common bile duct.

BACKGROUND: Several systemic factors, including jaundice, long-term corticosteroid therapy, diabetes and malnutrition, increase the risk of anastomotic dehiscence. The local application of molgramostim (recombinant human granulocyte-macrophage colony stimulating factor) has been reported to improve impaired dermal wound healing. Since jaundice, one of the systemic risk factors for anastomotic dehiscence, causes significant impairment of anastomotic healing, we hypothesized that locally injected molgramostim could improve the healing of bowel anastomoses in bile-duct-ligated rats used as an experimental model for jaundice. METHODS: Eighty-six Sprague-Dawley rats were randomized into 4 groups of 20-22 animals each as follows: group 1--colonic anastomosis only; group 2--laparotomy followed 7 days later by colonic anastomosis; group 3--common-bile-duct ligation (CBDL) followed 7 days later by colonic anastomosis (control group); group 4--CBDL followed by colonic anastomosis with locally applied molgramostim. Laparotomy was performed under anesthesia in group 2 rats. In groups 3 and 4, laparotomy was followed by ligation and dissection of the common bile duct. After 7 days, colonic anastomosis was performed; in group 4 rats, molgramostim (50 microg) was injected into the perianastomotic area. On postoperative day 3, rats were killed, and the bursting pressures and hydroxyproline levels measured. Two rats from each group were selected for histopathological examination. RESULTS: The mean bursting pressure in group 4 was significantly higher than that in group 3 (37.8 v. 30.5 mm Hg [p < 0.01]). The mean hydroxyproline level in group 3 was significantly lower than that of the other groups (2.7 v. 3.1-3.5 mg/g tissue [p < 0.01]). On histopathological examination, specimens from group 4 rats showed an increased mononuclear cell population and a smaller gap on the anastomotic line than those from group 3. CONCLUSION: The local injection of molgramostim improves healing of the impaired wound in rats subjected to CBDL.     

Effect of sildenafil citrate on an orthotopic prostate cancer growth and metastasis model

PURPOSE: We characterized the effects of sildenafil citrate on the growth and metastasis of human prostate cancer cells in nude mice. MATERIALS AND METHODS: The androgen independent human prostate cancer cell line PC-3 was inoculated into the prostate of nude mice to produce orthotopic primary prostate cancers and metastases. Sildenafil citrate gavage was started on day 31 after tumor cell inoculation and given every other day 15 times (30 days). The 7 mice in the low dose group received 25 mg/kg body weight sildenafil citrate per gavage, while the 7 in the high dose group received 50 mg/kg body weight sildenafil citrate and the 9 in the control group received water. Autopsy was performed on day 75 to evaluate primary tumor growth and metastasis. Plasma cyclic guanosine monophosphate concentrations were measured after the single dose of 50 mg/kg sildenafil citrate in the mice. RESULTS: Plasma cyclic guanosine monophosphate concentration increased 4-fold 1 hour after sildenafil citrate administration. The plasma concentration decreased rapidly and returned to normal after 8 hours. There was no significant difference in tumor weight between any of the 3 groups. The number of metastatic lymph nodes correlated significantly with primary tumor weight (p = 0.03) with a correlation coefficient of 0.454 but there was no significant correlation between the number of involved lymph nodes and sildenafil administration. Distant metastases were not significantly promoted by sildenafil administration. CONCLUSIONS: Incontinuous oral administration of sildenafil citrate did not promote primary tumor growth and metastasis in an orthotopic prostate cancer model.     

Sildenafil citrate inhibits agonist induced contractions in isolated rat seminal vesicles

PURPOSE: Sildenafil is reported to regulate smooth muscle contractility through nitric oxide-cyclic guanosine monophosphate, not only in the corpus cavernosum. Its possible effects on seminal vesicle contractility might be of importance with respect to premature ejaculation. We investigated the effects of sildenafil citrate (Pfizer, New York, New York) on agonist induced isometric contractions of the rat seminal vesicle in vitro. MATERIALS AND METHODS: Seminal vesicles isolated from adult male Wistar rats were suspended in an organ bath and contracted by NE (10 microM), ACh (10 microM) or KCl (60 mM) (Sigma, Deisenhofen, Germany). The effects of sildenafil citrate (100 to 300 microM) were evaluated in terms of mean contraction amplitude, the area under force-time curves and isometric contractility indexes. RESULTS: Sildenafil citrate (300 microM) significantly inhibited the mean amplitude +/- SEM of contractile responses induced by NE (1,061 +/- 153 vs 271 +/- 65 mg, p <0.0001), ACh (475 +/- 51 vs 68 +/- 17 mg, p <0.0001) and KCl (546 +/- 71 vs 59 +/- 18 mg, p <0.0002). It also caused dose dependent concomitant decreases in the area under force-time curves. Additionally, pretreatment with sildenafil citrate markedly prevented the contractile response to NE, ACh and KCl. CONCLUSIONS: Our results indicate that sildenafil citrate inhibits the contractions of isolated rat seminal vesicle that are induced by NE, ACh or KCl. Future studies may support an in vivo effect of sildenafil for delaying or inhibiting seminal vesicle emission, thereby, promoting improvement in patients with premature ejaculation.     

Effects of pneumoperitoneum with or without colostomy on rat colonic anastomotic healing

Background: Elevated intra‐abdominal pressure and colostomy have adverse effects on colonic anastomoses. The aim of the present study was to investigate the effects of laparoscopic colon surgery with and without diverting colostomy on healing of colonic anastomoses in an experimental model. Methods: Thirty‐six male rats were divided into three equal groups: group 1, control (colonic anastomosis and anaesthesia for 180 min only); group 2, 180 min pneumoperitoneum and colonic anastomosis; and group 3, similar to group 2 with a proximal colostomy. On day 7, bursting pressures, tissue hydroxyproline and nitric oxide concentrations and histopathological inflammation scores were determined and compared. Results: Mean bursting pressures were higher in the control group than the two pneumoperitoneum groups (P = 0.0003). Mean tissue hydroxyproline concentrations showed no difference (P > 0.05). Mean tissue nitric oxide concentrations were significantly increased in the control group (P = 0.0013). Histopathological scores demonstrated increased inflammatory response in group 3 compared to the controls (P = 0.0009). Conclusion: Pneumoperitoneum delays collagen maturation and impairs anastomotic strength in the colon. Following pneumoperitoneum, performance of a diverting loop colostomy to protect the anastomosis will not have additional detrimental effects on anastomotic healing.     

Effect of sildenafil citrate in nicotine-induced ischemia: An experimental study using a rat model

Recent experimental and clinical studies have demonstrated the negative effects of nicotine on the viability of skin flaps. Necrotic damage to skin flaps can result in significant complications including delayed wound healing, dehiscence and wound contraction. Phosphodiesterase type 5 inhibitors, such as sildenafil citrate, have a protective effect in ischemic injuries of the brain, kidney, myocardium, spinal cord, ileum and testes. In the present study, the authors evaluated the effect of sildenafil citrate on the viability of skin exposed to nicotine-induced ischemia in Sprague Dawley rats. In the preoperative period, the rats were divided into three groups of 10 rats each. Group C was treated with subcutaneous saline and group S and group N were treated with 2 mg/kg nicotine, administered subcutaneously twice per day for 28 days. McFarlane flaps were created in all experimental animals using an incision measuring 7 cm × 3 cm. Postoperative treatment varied among the groups: group S was treated with 20 mg/kg/day sildenafil citrate, while group C and group N were treated with equivalent doses of saline for seven days. A laser Doppler flow meter was used to monitor the microvasculature. Preoperative measurements of the microvasculature revealed decreased blood flow in group N and group S, both of which were treated with subcutaneous nicotine. During the postoperative evaluation, a trend toward increased blood flow was observed in group S compared with the group with nicotine-induced ischemia treated with saline alone postoperatively (group N). A visual fluorescein dye test was used to predict skin viability and demonstrated diminished skin viability in group N and group S (P<0.05) during the preoperative period. Following treatment with sildenafil for seven days, a statically significant improvement in skin viability was observed in group S (P<0.05). Nicotine decreased blood flow within the skin and impaired skin viability, while postoperative application of sildenafil significantly ameliorated the ischemic effects of nicotine and improved skin viability. Future studies will be required to evaluate the clinical use of sildenafil for the improvement of blood flow in ischemic injury of the skin.     

Dose-dependent protective effect of sildenafil citrate on testicular injury after torsion/detorsion in rats

This experiment was designed to investigate the effect of sildenafil citrate on testicular injury after unilateral testicular torsion/detorsion (T/D). Thirty-seven adult male Wistar albino rats were divided into four groups: sham operated group (group 1), T/D+ saline (group 2), T/D+ 0.7 mg sildenafil citrate (group 3) and T/D+ 1.4 mg sildenafil citrate (group 4). Testicular torsion was created by rotating the right testis 720° in a clockwise direction for 2 h in other groups, except for group 1, which was served as sham group. The level of GSH (P < 0.05) in the testis in the group 2 were significantly lower (P < 0.05) and the levels of MDA and NO (P < 0.01 for both) in the testis were significantly higher when compared with those of the group 1. Administration of low dose sildenafil citrate prevented the increases in MDA and NO levels and decreases in GSH values induced by testicular torsion. However, administration of high dose sildenafil citrate did not have any effect on these testicular tissue parameters (P > 0.05). Also, mean values of seminiferous tubules diameters, germinal cell layer thicknesses and mean testicular biopsy score were significantly better in group 3 than groups 2 and 4. These results suggest that T/D injury occurred in testis after unilateral testicular T/D and that administration of low dose sildenafil citrate before detorsion prevents ischemia/reperfusion cellular damage in testicular torsion. Sildenafil citrate probably acts through reduction of reactive oxygen species and support antioxidant enzyme systems.     

Methylene blue prevents surgery-induced peritoneal adhesions but impairs the early phase of anastomotic wound healing.

OBJECTIVES: Adhesion formation continues to be an important problem in gastrointestinal surgery. In recent years, methylene blue (MB) has been reported to be an effective agent for preventing peritoneal adhesions. However, its effects on the wound healing process are unknown. In the present study, we investigated the effects of MB on the early and late phases of anastomotic wound healing and on adhesion formation. METHODS: We randomly categorized 92 rats into 2 groups in bursting pressure measurements and 50 rats into 3 groups in the adhesion model. We divided the animals into saline-treated (n = 46) or MB-treated (n = 46) groups. Bursting pressures of the anastomoses were measured on postoperative days 3 and 7. In biochemical studies, tissue hydroxyproline levels, total nitrite/nitrate levels and nitric oxide synthase activity were measured on postoperative days 3 and 7. In the adhesion model, we randomly categorized rats into sham (n = 10), saline-treated (n = 20) and MB-treated (n = 20) groups, and the formation of intraperitoneal adhesions was scored on postoperative day 14. We compared the measurement of bursting pressure and biochemical measurements of tissue hydroxyproline levels, total nitrite/nitrate levels and nitric oxide synthase activity. Histopathological findings of specimens were presented. RESULTS: During the early phase of wound healing (postoperative day 3), bursting pressures, tissue hydroxyproline, total nitrite/nitrate levels and nitric oxide synthase activity in the MB-treated group were significantly lower than those of the saline-treated group. On postoperative day 7, there was no significant difference in these parameters between MB and saline-treated groups. In the adhesion model, MB caused a significant reduction in the formation of peritoneal adhesions. CONCLUSION: MB prevents peritoneal adhesions but causes a significant impairment of anastomotic bursting pressure during the early phase of the wound healing process by its transient inhibitory effect on the nitric oxide pathway.     

Assessment of heme oxygenase-1 （HO-1） activity in the cavernous tissues of sildenafil citrate-treated rats

Aim： To assess heine oxygenase-1 （HO-1） activity in the cavemous tissue of sildenafil citrate-treated rats. Methods： One hundred and ninety-two Sprague-Dawley male rats, divided into four equal groups, were investigated. Group 1, the control group, received regular animal chow; group 2 received sildenafil citrate by intragastric tube; group 3 received sildenafil and HO inhibitor （zinc protoporphyrin, ZnPP）; and group 4 received sildenafil and nitric oxide synthase （NOS） inhibitor L-nitroarginine methyl ester （L-NAME）. Twelve rats from each group were killed after 0.5 h, 1 h, 2 h and 3 h of drug administration. Then HO-1 activity, cGMP levels and NOS enzymatic activity in the cavernous tissues were estimated. Results： In cavemous tissue, HO-1 activity, NOS enzymatic activity and cGMP concentration increased significantly in sildenafil-treated rats compared to other groups throughout the experiment. Rats receiving either HO or NOS inhibitors showed a significant decrease in these parameters. HO- 1 cavemous tissue activity and NOS enzymatic activity demonstrated a positive significant correlation with cGMP levels （r = 0.646, r = 0.612 respectively; P 〈 0.001）. Conclusion： The actions of PDE5 inhibitor sildenafil citrate in the cavernous tissue are partly mediated through the interdependent relationship between both HO-1 and NOS activities.     

The influence of sildenafil on random skin flap survival in rats: an experimental study.

Sildenafil (Viagra), a selective and specific inhibitor of cyclic guanosine monophosphate (cGMP) phosphodiesterases (PDEs), is currently marketed for the treatment of erectile dysfunction. Sildenafil is a potent and highly selective PDE-V inhibitor and enhances smooth muscle relaxation in human. Systemic arterial and venous smooth muscle cells contain PDE-V and nitric oxide (NO) which is a major mediator of relaxation of the smooth muscle cell. The aim of the present study is to investigate, in a rat model, the potential effect of sildenafil on survival of random pattern skin flaps. For this purpose, 32 Sprague-Dawley rats were used and a McFarlane-type caudally based skin flap was designed on the dorsum of the rat (2.5 x 8 cm). Rats were divided into four groups: One control (Group D), and three treatment groups (Groups A, B, C). Sildenafil was administered orally to the experiment groups; Group A: 3 mg/kg/single dose a day, Group B: 10 mg/kg/single dose a day and Group C: 10 mg/kg/twice dose a day. The areas of flap necrosis were measured in each group. The extent of viable flap areas were expressed as a percentage of total flap area, and differences were studied by Completely Randomised Experimental design. The areas of necrosis of skin flaps decreased depending on sildenafil dose, but viability of the flaps treated with 3 mg/kg/day was not different than the control group. The flaps receiving 2 x 10 mg/kg/day sildenafil gave the highest (P < 0.01) survival rate. As a conclusion, sildenafil may have a dose dependent effect to increase flap survival in random skin flaps.     

Peritoneal release of TNFalpha and IL-6 after elective colorectal surgery and anastomotic leakage.

The evaluation of postoperative peritoneal drainage fluid tumor necrosis factor (TNF)alpha and interleukin (IL)-6 was studied prospectively over a 7-day period in 25 patients operated on for neoplastic colorectal diseases. In 22 cases, colon or rectum carcinoma was the reason for surgery, and in 3 patients resection was performed because of colonic adenoma. All patients received either an end-to-end colo-colonic or colorectal anastomosis. Of this group, 22 patients were free of complications defined as uneventful postoperative course without any signs of anastomotic leakage until the 14th postoperative day. All of these patients showed a significant rise in peritoneal TNFalpha with maximum on the 7th day during the study period (p <.05). In contrast, peritoneal IL-6 levels remained constant without significant change in time (p >.05). Three patients underwent relaparotomy because of anastomotic leakage. In these patients, peritoneal TNFalpha concentrations showed a rise until the day of operative confirmation of anastomotic leakage. This rise preceded the day of operative confirmation by at least 1 day but did not change significantly in time (p =.59). Peritoneal IL-6 concentrations in patients with anastomotic leakage remained constant and also did not change significantly in time (p =.21). After elective colorectal surgery, neither postoperative abdominal drainage fluid TNFalpha nor IL-6 monitoring is helpful to decide on the need for revision in patients with anastomotic leakage.     

Effects of Cholerella sp. microalgae extract on colonic anastomosis in rats with protein–energy malnutrition

Objective  Algae, which are used as supplementary nutrients in various countries, are products rich in protein, vitamins and minerals. The aim of this study was to investigate the effects of algae extracts on the healing of colonic anastomosis in malnourished rats. Method  Seventy‐two rats were randomized to three groups. Group 1 was fed with standard diet for 15 days, before and after the colonic anastomosis. Groups 2 and 3 were fed with a malnutrition diet for 15 days prior to colonic anastomosis and then with the basic diet for 15 days there after. Group 3 also received an extract of algae derived from Cholerella sp. via oral gavage postoperatively, in addition to the basic diet. Rats were killed on the 3rd, 7th and 15th postoperative day. Blood samples were collected to evaluate prealbumin, transferring and albumin levels. Anastomotic bursting pressures (BPs), histopathology and tissue hydroxyproline levels were evaluated after killing. Results  In group 3, the prealbumin level on the 3rd postoperative day and transferrin and albumin levels on the 7th and 15th postoperative days were significantly increased compared with the other groups (P < 0.05). Tissue hydroxyproline levels and anastomotic BPs of group 3 were significantly higher than in group 2 on the 3rd, 7th and 15th postoperative days (P < 0.05). Histopathological examination of the anastomosis revealed significantly better healing patterns for group 3 than for groups 1 and 2 (P < 0.05). Conclusion  Extract derived from Cholerella sp. microalgae has favourable effects on healing of experimental colon anastomoses.     

Effects of CO2 Pneumoperitoneum on Anastomotic Healing in Rats Receiving Preoperative 5-Fluorouracil Neoadjuvant Chemotherapy

ABSTRACT?

Background: When used separately, antineoplastic agents and carbon dioxide (CO₂) pneumoperitoneum have been reported to impair anastomotic healing in experimental animals. However, the effects of their combined use have not been previously investigated. The aim of this study was to investigate the possibility that neoadjuvant chemotherapy with 5-fluorouracil followed by CO₂ pneumoperitoneum would affect the healing of anastomoses in the colon. Methods: Sprague-Dawley rats (n = 48) were given 5-fluorouracil (20 mg/kg/day) for 5 days, and were then assigned to one of the three groups. Prior to surgery, the control group received no pneumoperitoneum. The other two groups received pneumoperitoneum at 6 and 12 mmHg, respectively, for 2 hr. The large intestine was transected and anastomosis was performed via median laparotomy. On postoperative days 3 and 7, relaparotomy was performed in half of the rats in each group. From the colon, a segment including the anastomosis was excised. Tissue hydroxyproline levels were measured. For histological evaluation, the Verhofstad scale was modified and used. Results: No significant differences in hydroxyproline levels were seen across the groups on postoperative days 3 or 7. However, by postoperative day 7, polymorphonuclear leukocytes and necrosis in the 6-mmHg group had decreased markedly, and granulation had improved. Conclusion: Overall, these findings suggest that preoperative 5-fluorouracil therapy followed by pneumoperitoneum at 6 or 12 mmHg does not impair anastomotic healing.     

Modulation of growth factor and cytokine expression by nitric oxide during rat colon anastomotic healing

We have previously shown that inhibition of nitric oxide generated by inducible nitric oxide synthase (iNOS) results in impaired colon anastomotic healing. Therefore, we proceeded to assess whether disruption of iNOS activity alters the normal pattern of growth factor expression during anastomotic healing. Two groups of male Sprague-Dawley rats underwent distal colonic division and anastomosis, jugular venous catheterization and subcutaneous placement of polyvinyl alcohol sponges. The first group (n = 10) received q8 hour intravenous injections of 10 mg/kg L-N-iminoethyl-lysine (L-NIL, a selective inhibitor of iNOS), while the second group (n = 12) received equal volumes of saline. On postoperative day 5, animals were sacrificed and anastomotic bursting pressure was determined. Histologic sections of the anastomosis were subjected to in situ hybridization versus mRNA of the proteins listed below. Positive controls were reacted with a poly-thymidine (poly-T) probe versus ubiquitous mRNA poly-adenine tails. Positively stained cells were quantified using a calibrated optical grid encompassing 0.5 mm² area centered over the anastomosis. Results are reported as the number of positive cells per 1000 cells positive for poly-T. L-NIL treated animals demonstrated an 18% decrease in wound fluid NOX compared to controls (29.2 ± 1.2 vs. 34.6 ± 2.0 iM, mean ± SEM; P = 0.035). This corresponded to a 17% decrease in anastomotic bursting pressure (153 ± 4 vs. 182 ± 8 mm Hg, mean ± SEM; P < 0.05). L-NIL also markedly increased the number of cells expressing transforming growth factor-β, tumor necrosis factor-ct, vascular endothelial growth factor, and both inducible and endothelial forms of nitric oxide synthase. L-NIL had no effect on the expression of basic fibroblast growth factor. The data demonstrate that iNOS inhibition markedly disrupts the profile of cytokine and growth factor mRNA normally expressed during anastomotic healing. This provides in vivo evidence that NO modulates gene expression during anastomotic healing. Presented at the Forty-First Annual Meeting of The Society for Surgery of the Alimentary Tract, San Diego, California, May 21–24, 2000 (poster presentation). This work was supported by funding from grants NIH #GM54566(AB) and NIH #T32 DK07713-03.     

Chronic inhibition of nitric‐oxide synthase induces hypertension and erectile dysfunction in the rat that is not reversed by sildenafil

Study Type – Aetiology (case control)
Level of Evidence 3b

OBJECTIVE

To evaluate the effect of N(G)‐nitro‐l ‐arginine methyl ester (L‐NAME)‐induced hypertension (HT) on erectile function in the rat and determine if the phosphodiesterase (PDE)‐5 inhibitor, sildenafil, can reverse the effects of nitric oxide (NO) deficiency, as HT is a risk factor for erectile dysfunction (ED) and the NO synthase (NOS) inhibitor L‐NAME induces NO‐deficient HT.

MATERIALS AND METHODS

Thirty‐six adult Sprague‐Dawley male rats were divided into three groups, i.e. a control, L‐NAME‐HT (40 mg/rat/day in the drinking water for 4 weeks), and sildenafil‐treated L‐NAME‐HT (1.5 mg/rat/day sildenafil, by oral gavage concomitantly with L‐NAME). The erectile response expressed as a ratio of intracavernosal pressure (ICP)/mean arterial pressure (MAP), evaluated after electrical stimulation of the right cavernous nerve. The isometric tension of corpus cavernosum smooth muscle (CCSM) was measured in organ‐bath experiments. NOS expression was determined immunohistochemically for neuronal (n)NOS and by Western blot analysis for endothelial (e) and inducible (i) NOS protein. cGMP levels were evaluated by enzyme‐linked immunosorbent assay.

RESULTS

The erectile response was diminished in the HT group. Nitrergic and endothelium‐dependent relaxation was reduced, while the relaxation response to sodium nitroprusside and contractile response to phenylephrine were not altered in CCSM from L‐NAME‐treated rats. HT rats showed decreased expression of nNOS, whereas eNOS and iNOS protein expression was increased. Sildenafil partly restored endothelial and molecular changes in CCSM from HT rats, but did not reverse the decreased erectile response, even as cGMP levels returned to normal levels.

CONCLUSIONS

Sildenafil treatment did not correct the ED in L‐NAME‐treated HT rats. Under sustained high blood pressure, up‐regulation of PDE5 expression failed to reverse the depletion of neuronal NO and/or impaired nNOS activity. However, endothelium‐dependent relaxation was restored. Drug targeting of neuronal dysfunction might delay the onset of ED in HT.     

Effects of locally applied recombinant human granulocyte-macrophage colony-stimulating factor on ischemic bowel anastomoses in rat

BACKGROUND: Ischemia is one of the most common causes of anastomosis disruption. In the present study we investigated the effect of locally injected recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) on ischemic bowel anastomosis in rats. METHODS: 144 male Sprague-Dawley rats were randomized into four groups: in group 1, colon anastomoses were performed; in group 2, anastomoses were performed then 50 microg rhGM-CSF was injected subserosally into the perianastomotic area; in group 3, anastomoses were performed on ischemic colon segments, and in group 4, colon anastomoses were performed on ischemic segments and then 50 microg rhGM-CSF was injected subserosally into the perianastomotic area. On the 3rd and 7th post-operative days, the rats were sacrificed and anastomotic bursting pressures were measured. Hydroxyproline contents were studied on the tissues from the anastomotic line. Three anastomotic segments were saved from each group for histopathological studies before bursting pressure measurement. RESULTS: The bursting pressure in group 3 was significantly weaker than in the other groups. There were no significant differences between the bursting pressures in groups 1 and 4. The levels of hydroxyproline content in group 4 were significantly greater than in group 3. CONCLUSIONS: These data suggest that the local injection of rhGM-CSF improves the healing of ischemic and even normal colon anastomoses.