Vascular anatomy of the small intestine-a comparative anatomic study on humans and pigs.

BACKGROUND: Porcine models are well established for studying intestinal anastomotic healing. In this study, we aimed to clarify the anatomic differences between human and porcine small intestines. Additionally, we investigated the influences of longitudinal and circular sutures on human small intestine perfusion. METHODS: Intestines were obtained from human cadavers (n = 8; small intestine, n = 51) and from pigs (n = 10; small intestine, n = 60). Vascularization was visualized with mennige gelatin perfusion and high-resolution mammography. Endothelial cell density was analyzed with immunohistochemistry and factor VIII antibodies. We also investigated the influence of suture techniques (circular anastomoses, n = 19; longitudinal sutures, n = 15) on vascular perfusion. RESULTS: Only human samples showed branching of mesenteric vessels. Compared to the pig, human vessels showed closer connections at the entrance to the bowel wall (p = 0.045) and higher numbers of intramural anastomoses (p < 0.001). Porcine main vessels formed in multifilament-like vessel bundles and displayed few intramural vessel anastomoses. Circular anastomoses induced a circular perfusion defect at the bowel wall; longitudinal anastomoses induced significantly smaller perfusion defects (p < 0.001). Both species showed higher vascular density in the jejunum than in the ileum (p < 0.001). Human samples showed similar vascular density within the jejunum (p = 0.583) and higher density in the ileum (p < 0.001) compared to pig samples. CONCLUSION: The results showed significant differences between human and porcine intestines. The porcine model remains the standard for studies on anastomotic healing because it is currently the only viable model for studying anastomosis and wound healing. Nevertheless, scientific interpretations must consider the anatomic differences between humans and porcine intestines.

Balancing zinc deficiency leads to an improved healing of colon anastomosis in rats.

BACKGROUND: In colorectal surgery, anastomotic leakage is a relevant complication. The aim of this study was to investigate whether intraperitoneally (i.p.) administered zinc improves the healing of colon anastomosis in rats. MATERIALS AND METHODS: Male Wistar rats (66) received zinc-deficient diet for 21 days. To determine the effective dose of zinc which is necessary to compensate this deficiency, preliminary analysis in 30 rats were performed. In these rats, analysis by atom-absorption spectrophotometry revealed a dose of 1.0 mg zinc aspartate/kg body weight to be the compensatory dosage. In the remaining zinc-deficient rats (n= 36), a transverse colonic anastomosis was performed. Eighteen rats received either a zinc supplementation i.p. or 0.9% NaCl i.p. (n = 18; control group). On postoperative days 3, 5, and 14, the surface of the mucosal villi, expression of MMP 2, MMP 8, MMP 13, TIMP 1, as well as the collagen types I/III ratio were analyzed. RESULTS: Protein expression of MMP 2 and MMP 8 was significantly higher in the anastomosis of the zinc group on day 3 and on day 5. The collagen types I/III ratio was significantly increased in the zinc group on days 5 and 14. CONCLUSION: Balancing zinc deficiency benefits wound healing of colonic anastomosis qualitatively due to an increased collagen type I/III ratio. Surprisingly, these zinc supplements, however, increased the expression of MMP 2 and MMP 8 that are supposed to impair wound healing in case of an over-expression. Thus, further investigations are needed to elucidate the influence of zinc supplementation on regulation of MMPs.

Erythropoietin (EPO) influences colonic anastomotic healing in a rat model by modulating collagen metabolism.

INTRODUCTION: Anastomotic failure is one of the most frequent complications in abdominal surgery. During anastomotic healing. the strength of the intestinal tissue nearby is closely related to the accumulation of collagen in interlinked scar tissue. This in turn is influenced, among other things, by single groups of matrixmetalloproteinases, especially collagenases (MMP-1, -8, and -13) and gelatinases (MMP-2 and -9). EPO is known to induce the expression of tissue-inhibitor-of-matrixmetalloproteinases-1 (TIMP-1) and thereby to down-regulate MMPs. MATERIALS AND METHODS: We used a rat as an experimental model and applied a high dose of EPO (5U/g BW s.c.), one dose 24 h before operation (as pre-conditioning) and one dose directly after performing a colonic anastomosis. After 3 and after 5 d, respectively, immunohistochemical stainings for MMP-2, -8, and -9 as well as TIMP-1 were carried out and evaluated semiquantitatively for each layer of the colonic wall. Sirius-red staining and cross-polarization microscopy were evaluated and the collagen I/III ratio calculated. Anastomotic and colonic tissue distal to the anastomosis were used to determine collagen content. RESULTS: We found increased bursting pressure 5 d post-surgery after applying erythropoietin. It was thus shown that EPO influences collagen metabolism and changes the collagen I/III ratio in the colon distal to the anastomosis. The evaluation of immunohistochemistry did not show the expected ubiquitous up-regulation of TIMP-1 and down-regulation of MMPs. Nevertheless, correlations between TIMP-1, MMP-8, and collagen I/III ratio could only be established after the application of EPO. CONCLUSION: Contrary to our hypothesis, the picture of TIMP-1 and of the regulation of the MMPs after the application of EPO is not as clear as expected. EPO improves anastomotic bursting strength and the correlation of TIMP-1, MMP-8, and collagen type I/III ratio can only be seen after the application of EPO.

Zinc deficiency impairs wound healing of colon anastomosis in rats.

BACKGROUND: Anastomotic leakage is a relevant surgical complication. The aim of the study was to investigate the influence of a controlled preoperative zinc deficiency on the extracellular matrix composition of colon anastomosis. MATERIALS AND METHODS: Forty male Wistar rats were randomized to either a zinc deficiency group (n = 20) or a control group (n = 20). In each animal, a transverse colonic end-to-end anastomosis was performed. On postoperative day 7, the surface of the mucosal villi, expression of matrix metalloproteinases (MMP) 2, 8, 9, and 13, and both the number of proliferating cells (Ki67) and apoptotic cells, as well as the collagen types I/III ratio were analyzed. Within the anastomotic area the mesenterial region and the antimesenterial region were analyzed separately. RESULTS: In each group, one anastomotic leakage was detected. Expression of both MMP 2, 9, and 13 was significantly higher, and expression of Ki67 was significantly reduced in the zinc deficient group both mesenterial and antimesenterial. The collagen types I/III ratio was reduced in the zinc deficiency group by trend, without statistical significance neither mesenterial nor antimesenterial. Likewise, zinc deficiency affected neither the expression of MMP 8 nor the rate of apoptotic cells, respectively. Analyses of the surface of the mucosal villi revealed no significant differences comparing the groups with neither mesenterial nor antimesenterial. CONCLUSIONS: Our study constitutes the known negative effect of zinc deficiency on wound healing. Zinc deficiency significantly increased the activity of MMPs (2, 9, and 13), caused a reduced collagen type I/III ratio, and delayed cell proliferation and quality of intestinal wound healing.

Intraperitoneally applied gentamicin increases collagen content and mechanical stability of colon anastomosis in rats.

BACKGROUND: Anastomotic leakage remains a serious complication in colorectal surgery, and is being caused by a multitude of factors. Recent reports reveal changes of the extracellular matrix as risk factors as well as gentamicin as a potential agent to influence wound healing. This experimental study was initiated to investigate the influence of intraperitoneally applied gentamicin on colonic anastomotic wound healing and in particular on mechanical stability, overall collagen content and collagen type I/III ratio. MATERIALS AND METHODS: Sixty Sprague Dawley rats were randomized to one of two groups. In each animal, a standard transverse colonic end-to-end anastomosis was performed. Immediately postoperative, either 5 ml gentamicin (1 ml/kg bodyweight) or NaCl 0.9% was applied intraperitoneally. On postoperative days 3, 5, and 14, ten of the animals in each group were sacrificed. Measurements of the anastomosis bursting pressure were performed on postoperative days 3 and 5. At each explantation time, the collagen per protein ratio, the collagen types I/III ratio, and both the expression of MMP-2, -9, and Ki67 were analyzed. RESULTS: None of the animals died. None of the rats exhibited clinical evidence of anastomotic leakage. The bursting strength in the gentamicin group was significantly elevated on postoperative day 5. Both the overall collagen content and the collagen type I/III ratio in the gentamicin group were significantly increased 3, 5, and 14 days postoperatively compared to the control group. The expression of MMP-9 was significantly elevated in the gentamicin group both 3 and 5 days postoperatively. In contrast, there were no significant differences in the expression of MMP-9 14 days postoperatively. All investigated samples demonstrated positive staining for MMP-2 and Ki67 without statistically significant differences at any term, respectively. CONCLUSIONS: The present data confirm that intraperitoneally applied gentamicin is able to enhance healing and stability of colonic anastomosis due to an increase of both the overall collagen content and collagen type I/III ratio.

Is there anybody out there: what do senior surgeons expect of their youngsters?

Surgery is indeed one of the most fascinating medical professions. However, it is also a stressful field of work with a high workload, and often leaves little time for personal and family needs. Within the last decade, a noticeable decline occurred in the willingness of medical students to enter a surgical residency. In fact, Generation Y is highly interested in a medical career with a respectful working atmosphere and balanced work and private life, as published in several recent papers. Therefore, surgery must evolve with the times to retain its attractiveness as a career choice for medical students and to compete for the best talents from Generation Y. However, little is known about what senior surgeons really expect from young surgical residents. On the basis of a recent survey by the Professional Association of German Surgeons, this paper tries to give some insights in this very relevant topic and a perspective on how to increase the attractiveness of our fascinating specialty. In fact, in this survey, senior surgeons defined a very clear requirement profile for surgical residency applicants. While the colleagues defined accurate applicant documents, a previous internship, self-motivation, and impressions from the job interview as the most important factors for a successful application for a surgical residency, a standard period of study or a dissertation was deemed of lower importance.

Collagen in colon disease.

The pathophysiology of wound healing in the bowel wall suggests that collagen and matrix metalloproteinases (MMPs) have an important role in the changes of the bowel wall seen in several colonic diseases. Several recent studies suggest that disturbances of the collagen texture and the extracellular matrix (ECM) metabolism are major factors leading to the onset of diverticular disease. Changes of the ECM also play a role in the development of inflammatory bowel diseases. Regarding the permanent remodeling of the bowel wall, any imbalance of the ECM could support the onset of chronic inflammation and the development of fistula formation, such as that seen in patients with Crohn's disease. Disturbances of the ECM play a role in the pathogenesis of anastomotic leakage after large bowel surgery and suggest the presence of a genetically defined risk population with disturbed wound healing mechanisms. This concept could explain the well known situation where an anastomotic breakdown is observed, despite the absence of other known risk factors and after a technically correct anastomosis.

Gentamicin supplementation of polyvinylidenfluoride mesh materials for infection prophylaxis.

Hernia repair evolved from pure tissue repair to mesh repair due to decreased recurrence rates. However, concern exists about mesh-related infections occurring even several years after initial operation. Therefore, a polyvinylidenfluoride (PVDF) mesh material was constructed and surface modified by plasma-induced graft polymerization of acrylic acid (PVDF+PAAc). Antimicrobial treatment was sought by binding of gentamicin (PVDF+PAAc+Gentamicin). In vitro efficacy and cytotoxicity was measured by agar diffusion test, L929 cytotoxicity testing and by analyzing the amount of gentamicin release from the mesh surface. In vivo biocompatibility was evaluated in 45 Sprague-Dawley rats. 7, 21 and 90 days after mesh implantation the amount of inflammatory and connective tissue as well as the percentage of proliferating (Ki67) and apoptotic cells (TUNEL) were analyzed at the perifilamentary region. Agar diffusion tests showed sufficient local antimicrobiotic effects against the bacteria tested after 24h of incubation. No signs of cytotoxicity could be identified by L929 testing. Furthermore, surface modification did not affect the in vivo biocompatibility. At the end of the observation period, no significant differences were found for the perifilamentary amount of inflammatory cells and connective tissue and the percentage of Ki67 and TUNEL positive stained cells. The presented data confirm that an antibiotic surface modification of PVDF mesh samples is feasible. By analyzing cytotoxicity in vitro as well as biocompatibility in vivo no side effects were observed.

Changes of the extracellular matrix as a risk factor for anastomotic leakage after large bowel surgery.

BACKGROUND: Despite improved surgical techniques, anastomotic leakage remains as a serious complication in colorectal surgery, producing increased morbidity and mortality. This prospective study was initiated to test the hypothesis that preexisting disorders in the extracellular matrix (ECM) may be a factor influencing the onset of anastomotic wound healing complications. METHODS: In this prospective study of 119 patients with colorectal anastomoses, 30 clinical parameters with possible influence on anastomotic complications were evaluated. From all patients, samples of macroscopically intact colonic tissue were obtained at the index operation. Crosspolarization microscopy was performed to analyze the collagen type I/III ratio, and immunohistochemical studies were done to determine the expression of matrix metalloproteinase (MMP) 1, 2, 9, and 13. The patients with uncomplicated postoperative healing were compared with those developing anastomotic leakage. RESULTS: Patients with impaired anastomotic healing exhibited a significantly lower collagen type I/III ratio compared with the controls. Significantly higher expression of MMP-1 and MMP-2 in the mucosal layers and of MMP-2 and MMP-9 in the submucosal layers was found in the normal bowel wall of the leakage group. These findings were statistically independent from the clinical parameters. CONCLUSION: The present study confirms the hypothesis that disturbances of the ECM play a role in the pathogenesis of anastomotic leakage after large bowel surgery.

The influence of inguinal mesh repair on the spermatic cord: a pilot study in the rabbit.

The permanent implantation of a polypropylene mesh during inguinal hernia repair causes chronic inflammatory changes in the surrounding tissue. We investigated the effect of this foreign body reaction on the structures of the spermatic cord in the rabbit. Eight Chinchilla rabbits underwent unilateral inguinal hernia repair by the Lichtenstein technique using Marlex (n = 4) or Ultrapro (n = 4) mesh. The contralateral side was operated upon using the Shouldice repair. Three animals served as controls. Three months after operation we analyzed testicular size, testicular temperature, and arterial perfusion by excitation light of a 780-nm laser after injection of 0.5 mg/kg indocyanin green. Histological evaluation included spermatogenesis (Johnsen score) and foreign-body reaction. Testicular volume increased about 10% after each operation. The decrease of arterial perfusion and testicular temperature was more significant after mesh repair than following Shouldice operation. After mesh implantation we found fewer seminiferous tubules classified as Johnsen 10 (Marlex: 51.3%, Ultrapro: 45.0%) than after Shouldice repair (63.8%) or in the controls (65.8%). The spermatic cord showed a typical foreign-body reaction at the interface between mesh and surrounding tissue, which was not detectable after Shouldice repair. Preserved cremasteric muscle fibers protected the structures of the spermatic cord. The inflammatory foreign-body reaction of the surrounding tissue induced by the inguinal prosthetic mesh includes the structures of the spermatic cord. This may have an influence also on spermatogenesis. Therefore, we recommend strict indications for implantation of a prosthetic mesh during inguinal hernia repair.

Reduced expression of collagen type I and increased expression of matrix metalloproteinases 1 in patients with Crohn's disease.

Crohn's disease (CD) is a chronic inflammatory bowel disease of still unknown etiology. The aim of our study was to find out whether there are any changes in the colonic wall of CD patients that could give hints for a predisposing disorder concerning the extracellular matrix, especially the collagen metabolism. Eight samples of colonic tissue from patients with Crohn's disease were compared to 14 specimens from patients without Crohn's disease. We performed a sirius red test for the overall collagen content and immunohistochemical studies examining differentiation between" collagen type I and type III and the expression of MMP-1 and MMP-13. In the bowel sections of patients with Crohn's disease, decreased levels of mature collagen type I with a resulting lower ratio of collagen I/III compared to patients without Crohn's disease were found (1.12 +/- 0.29 vs. 1.59 i 0.31). The expression of MMP-1 was significantly increased in the CD group (9.21 i 6.02 vs.6.02 i 1.98), whereas expression of MMP-13 showed no difference in both groups. Our study gives the first indication that preexisting changes of the extracellular matrix in the colonic wall may play a role in the pathogenesis of CD. Further studies have to be done to elucidate these interesting aspect of the pathogenesis in Crohn's disease.

Missing effects of zinc in a porcine model of recurrent endotoxemia.

BACKGROUND: Chronic human sepsis often is characterised by the compensatory anti-inflammatory response syndrome (CARS). During CARS, anti-inflammatory cytokines depress the inflammatory response leading to secondary and opportunistic infections. Proved in vitro as well as in vivo, zinc's pro-inflammatory effect might overcome this depression. METHODS: We used the model of porcine LPS-induced endotoxemia established by Klosterhalfen et al. 10 pigs were divided into two groups (n = 5). Endotoxemia was induced by recurrent intravenous LPS-application (1.0 microg/kg E. coli WO 111:B4) at hours 0, 5, and 12. At hour 10, each group received an intravenous treatment (group I = saline, group II = 5.0 mg/kg elementary zinc). Monitoring included hemodynamics, blood gas analysis, and the thermal dilution technique for the measurement of extravascular lung water and intrapulmonary shunt. Plasma concentrations of IL-6 and TNF-alpha were measured by ELISA. Morphology included weight of the lungs, width of the alveolar septae, and rate of paracentral liver necrosis. RESULTS: Zinc's application only trended to partly improve the pulmonary function. Compared to saline, significant differences were very rare. IL-6 and TNF-alpha were predominately measured higher in the zinc group. Again, significance was only reached sporadically. Hemodynamics and morphology revealed no significant differences at all. CONCLUSION: The application of zinc in this model of recurrent endotoxemia is feasible and without harmful effects. However, a protection or restoration of clinical relevance is not evident in our setting. The pulmonary function just trends to improve, cytokine liberation is only partly activated, hemodynamics and morphology were not influenced. Further pre-clinical studies have to define zinc's role as a therapeutic tool during CARS.

Deleterious effect of zinc in a pig model of acute endotoxemia.

By antiapoptotic effects and the induction of the heat-shock response, zinc is supposed to be a promising means of therapy during sepsis. As zinc also stimulates the expression of proinflammatory cytokines, its administration during the proinflammatory stage of septic shock might have adverse effects. Therefore, this study analyzes the influence of zinc during the acute phase of endotoxemia. In a pig model of acute endotoxemia, animals were divided into two groups: group I (n = 5) with saline treatment and group II (n = 5) with zinc treatment in close succession to lipopolysaccharide (LPS) (1.0 mu g/kg Escherichia coli endotoxin WO 111:B4). Hemodynamic and pulmonary monitoring was followed by combined reflection photometry, pulse oxymetry, blood gas samples, and temperature measurement. Plasma concentrations of tumor necrosis factor (TNF)alpha and interleukin (IL)-6 were analyzed by enzyme-linked immunosorbent assay (ELISA). Morphology included the weight of the lungs, the width of the alveolar septae, and the paracentral necrosis rate of the liver. After LPS infusion, group II (zinc) showed an impressive and significant deterioration of all pulmonary and most of the hemodynamical parameters compared to group I (saline). Levels of TNFalpha and IL-6 measured were significantly higher after zinc treatment. In accordance, we found significant more morphologic damages in group II (zinc). The almost simultaneous infusion of zinc and LPS complementary induced proinflammatory effects with a deleterious outcome. The same potentials characterizing zinc as a promising tool of prophylactic therapy in sepsis seem to ban its use during the acute phase.

Comparison of intestinal microcirculation and wound healing in a rat model.

PURPOSE: If a colonic or small bowel lesion is not accessible for endoscopic mucosal resection, enterotomy is a possible although rarely performed surgical technique. It has never been compared to circular anastomosis regarding leakage rate, bowel wall perfusion, and wound healing. Thus, the aim of this basic experimental scientific study was to investigate perianastomotic microcirculation and wound healing. METHODS: Forty rats were divided into four groups (1 jejunal anastomosis, 2 jejunal enterotomy, 3 colonic anastomosis, and 4 colonic enterotomy). Following anastomosis and enterotomy, the intestinal perfusion was measured using laser fluorescence angiography (IC-View). On postoperative day 7, the surface of the mucosal villi, expression of matrix metalloproteinases (MMP) 2, 8, 9, and 13, and the number of proliferating cells (Ki67) as well as the collagen types I/III ratio were analyzed. RESULTS: The perianastomotic microperfusion was significantly reduced in all groups compared to the reference region. The perianastomotic perfusion index was significantly reduced in group 1 compared with group 2, whereas the perfusion index in group 3 was slightly but not significantly reduced in comparison to group 4. Ki67 was elevated in both circular anastomosis groups. Surface of the mucosal villi, MMP expression, and collagen type I/III ratio revealed no significant differences. CONCLUSIONS: Our study affirms the theoretical consideration of a better microperfusion of the bowel wall following an antimesenterial enterotomy and demonstrates that enterotomy is not inferior compared to circular anastomosis. Even though enterotomy is a rarely used surgical technique, it should be regarded as a possible alternative in particular situations.

Suture-free small bowel anastomoses using collagen fleece covered with fibrin glue in pigs.

BACKGROUND: Several studies investigating anastomotic healing could objectify that the regularly used suture material leads to an impairment of wound healing due to ischemia at the anastomotic line. This study was initiated to test a hypothesis that a reduction of suture material leading to suture-free glued intestinal anastomoses is feasible and enables an improved wound healing. MATERIALS AND METHODS: Three different types of anastomoses were carried out at the small bowel of 16 pigs. Standard hand-sewn anastomoses, anastomoses with loose-fitting skin staples, and suture-free glued anastomoses using a fibrin covered collagen fleece. When the animals were killed, both gross inspection of the parietes, bursting pressure and tissue for histological study became the basis for evaluation. Analyses were also made regarding the collagen I/III ratio and the expression of MMP 1 and 13. RESULTS: Four leakages at the stapled, one at the sutured, and one at the glued anastomoses occurred. All other anastomoses healed without complications. The bursting pressure did not differ significantly between the groups. The macroscopic inspection and the microscopic examination both showed an improved healing pattern for the material reduced techniques without onset of a deep ulcer at the anastomotic line as seen at the conventional sutured anastomoses. These findings were supported by the immunohistochemical studies. CONCLUSIONS: These observations suggest that a suture-free bowel anastomoses using collagen fleece covered with fibrin glue is technically feasible. Obviously, a reduction of foreign body material at the anastomotic line avoids unnecessary ischemia and thus supports a physiological improved wound healing process.