Margin width is not predictive of residual disease on re-excision in breast conserving therapy.

BACKGROUND: There is lack of consensus regarding re-excision in breast-conserving therapy (BCT) and close margins. We hypothesize that margin width does not predict residual disease. METHODS: The cancer registry was queried from 2003 to 2008 for patients with BCT who underwent re-excision for <2-mm margins. Factors associated with additional disease were evaluated. RESULTS: One thousand eight hundred forty-three patients underwent BCT. Our re-excision rate was 42%. Clinicopathologic factors from 228 patients were analyzed. One hundred five patients (46%) had additional disease; of those, 58% had BCT and 42% mastectomy. One hundred twenty-three (54%) had no additional disease; of those 82% had BCT and 18% mastectomy. Of the 66 patients who underwent mastectomy, 44 (67%) had residual disease; of the 161 who had BCT, 61 (38%) had residual disease (P < 0.01). On univariate analysis, margin width did not correlate with residual disease. Multifocality, non-invasive histology, increasing number of close margins, and higher grade predicted additional disease (P < 0.05). On multivariate analysis, only number of close margins remained significant. CONCLUSIONS: Margin width does not predict additional disease. This challenges the practice of using this to select re-excision candidates. Our data suggest that tumor behavior and extent of disease, defined by volume of residual disease and invasiveness of histology, play a more significant role.

Diabetes-impaired healing and reduced wound nitric oxide synthesis: a possible pathophysiologic correlation.

BACKGROUND: Nitric oxide (NO) is synthesized in wounds, but its role in the healing process is not fully understood. The inhibition of NO production during wound healing is accompanied by decreased wound reparative collagen deposition. To further define the role of NO in reparative collagen accumulation, we studied its production during diabetes-induced wound healing impairment. METHODS: Male Sprague-Dawley rats (290 to 310 gm) were rendered diabetic by intraperitoneal streptozotocin administration. Seven days after induction of diabetes (blood glucose greater than 300 mg/dl), the rats underwent dorsal skin incision and subcutaneous implantation of polyvinyl alcohol sponges. Beginning on the day of wounding, 21 diabetic animals were treated with 3 units/day insulin via intraperitoneally implanted miniosmotic pumps. Ten days after injury, wound breaking strength was determined, and wound collagen accumulation and types I and III collagen gene expression were measured in subcutaneously implanted polyvinyl alcohol sponges. NO-synthesis, as measured by nitrite/nitrate accumulation, was determined in wound fluid and in supernatants of wound cell cultures. RESULTS: Streptozotocin-induced diabetes markedly impaired wound breaking strength and collagen deposition. A parallel decrease occurred in wound NO synthesis as reflected by decreased nitrite/nitrate concentration in wound fluid and in diminished ex vivo NO production by wound cells. Insulin treatment partially but significantly improved wound mechanical strength (p < 0.01) and collagen accumulation (p < 0.001). Decreased wound NO accumulation and ex vivo NO production by wound cells were also partially restored by insulin treatment. CONCLUSIONS: Impaired diabetic wound healing is paralleled by decreased wound NO synthesis, supporting the hypothesis that NO plays a significant role in wound reparative collagen accumulation.

Sepsis impairs anastomotic collagen gene expression and synthesis: a possible role for nitric oxide.

Although intra-abdominal sepsis is known to impair colon healing by inhibiting anastomotic collagen synthesis, the effect of systemic sepsis on this process is unknown. Endotoxins and cytokines associated with sepsis induce nitric oxide synthesis both systemically and locally within colonic tissue. We hypothesized that systemic sepsis impairs colonic healing and examined a possible correlation with nitric oxide expression. Male Sprague-Dawley rats received intraperitoneal injections of either saline (sham group) or Escherichia coli endotoxin (lipopolysaccharide 1 mg/100 g body weight) at Times -24 and -12 hr (LPS group). All animals underwent laparotomy and left colonic anastomosis at Time 0. At 24 and 96 hr postlaparotomy rats were sacrificed, the anastomoses excised, and [3H]-proline incorporation into protein measured as an index of total new protein synthesis (TNP). Digestion with purified collagenase yielded incorporation into the collagen fraction (CDP). Additional sham and LPS-treated rats were sacrificed at 24, 72, and 120 hr, the anastomoses excised, and nitric oxide synthase activity in the tissue measured by the conversion of [3H]-arginine to [3H]citrulline in an ex vivo culture system. Finally, sham and LPS rats were sacrificed at 120 hr for measurement of colon anastomotic bursting pressure. Systemic sepsis significantly impaired new collagen synthesis in anastomotic tissue at 24 hr compared to control samples (P < 0.02). No difference was noted at 96 hr. TNP synthesis was similar in both groups at 24 or 96 hr. Northern blot analysis confirmed a significant decrease in Type I and Type III collagen mRNA expression at 24 hr in septic rats. Anastomotic bursting pressure was also decreased in the septic group (P < 0.003). Sepsis elevated nitric oxide synthase activity in anastomotic tissue 24 hr postanastomosis, when compared to sham tissue (P < 0.0001). These data suggest that systemic endotoxin induces nitric oxide synthesis at the anastomotic site. The simultaneous dysregulation of collagen gene expression and synthesis with decreased anastomotic strength suggests a possible regulatory role for nitric oxide in gastrointestinal healing.

Acute protein-calorie malnutrition impairs wound healing: a possible role of decreased wound nitric oxide synthesis.

BACKGROUND: Nitric oxide is synthesized in wounds. Systemic inhibition of wound nitric oxide synthesis decreases wound collagen accumulation and wound mechanical strength. The role of nitric oxide during impaired healing is not known. In a model of impaired wound healing induced by acute protein-calorie malnutrition, we correlated wound healing parameters with wound nitric oxide synthesis. STUDY DESIGN: One group of Sprague-Dawley rats was rendered acutely malnourished by restricting its food intake to 50 percent of the food intake of an ad libitum-fed control group. Wound collagen accumulation and types I and III collagen gene expression were measured 10 days postwounding in subcutaneously implanted polyvinyl alcohol sponges. Nitric oxide synthesis was determined in wound fluid and in supernatants of wound cell cultures. RESULTS: Animals with acute protein-calorie malnutrition lost 10.4 +/- 0.8 percent, while controls gained 17.5 +/- 1.2 percent of their original body weight. Protein-calorie malnutrition reduced sponge hydroxyproline contents (995 +/- 84 compared with 1,580 +/- 109 micrograms/100 mg sponge, p < .001), indicating diminished wound collagen accumulation. Gene expression of type III, but not type I, collagen was decreased in wounds of protein-calorie malnutrition animals. Nitrite/nitrate and citrulline concentrations in wound fluid (p < .01) and in wound cell supernatants (p < .001) were also lower in protein-calorie malnutrition animals, indicating a net decrease in nitric oxide production. CONCLUSIONS: Impaired wound collagen accumulation caused by protein-calorie malnutrition may be a reflection of reduced nitric oxide synthesis within the wound.

Stimulation of fibroblast proliferation and matrix contraction by wound fluid.

Fibroblast proliferation and fibroblast-mediated matrix contraction are critical to wound healing. Different cytokines have been shown to modulate fibroblast functions but little is known about the physiological role of these soluble factors during wound repair. In these experiments we characterized a fibroblast stimulating factor in wound fluid. Wound fluid was obtained from subcutaneously implanted polyvinyl alcohol sponges harvested 10 days post-wounding (pool of 100 Lewis rats). Normal dermal fibroblasts were obtained from Lewis rats by an explant technique, while wound fibroblasts were isolated from sponges harvested 10 days post-wounding. Proliferation in response to 0.5% and 10% fetal bovine serum was assessed by [3H]-thymidine incorporation. A fibroblast-populated collagen lattice was used for assaying contractile properties. Wound fibroblasts demonstrated markedly diminished proliferative and enhanced contractile properties compared to normal dermal fibroblasts. 10% wound fluid (v/v) stimulated proliferation of normal dermal fibroblasts (119%, p < 0.001) and wound fibroblasts (103%, p < 0.001). Wound fluid also stimulated collagen gel contraction by normal dermal fibroblasts (24% at 24 hr and 16% at 72 hr, p < 0.01), but not by wound fibroblasts. Separation by Sephadex G-100 gel filtration identified the active factor in wound fluid to have a molecular weight of about 100 kDa. Characterization of the soluble factor showed it to be a protein (ammonium sulfate precipitation), sensitive to trypsin digestion, heat resistant (56 degrees C, 30 min), and neuraminidase resistant. The isoelectric point appeared to be 7.0, as determined by ion exchange chromatography. Mitogenic proliferation of thymic lymphocytes was not affected by the active factor, suggesting cell target specificity. These data demonstrate that the wound environment contains high molecular weight protein(s) that promote fibroblast functions, essential for the healing process.

Intra-abdominal sepsis impairs colonic reparative collagen synthesis.

BACKGROUND: Intra-abdominal infection is generally considered a contraindication to primary colon anastomosis. In order to elucidate the mechanisms by which sepsis affects colonic healing, we studied anastomotic new collagen and protein synthesis and collagen gene expression in a relevant animal model. METHODS: Forty male Sprague-Dawley rats (240 to 260 g) underwent sham laparotomy (SHAM, n = 18) or cecal ligation and single puncture (CLP, n = 22). After 24 hours, animals underwent single-layer left colon anastomosis. Animals were sacrificed either 1 or 4 days postanastomosis. Anastomotic segments of colon were excised, minced, and incubated with 4.5 muCi 3H-proline. After 3 hours, tissue 3H-proline incorporation was quantitated as an index of total new protein synthesis. The protein fraction was then digested with purified collagenase enzyme to determine 3H-proline incorporation into collagenase-digestible protein, an index of new collagen synthesis. Total RNA was extracted from anastomotic tissue samples and subjected to Northern blot analysis for type I and type III collagen genes. RESULTS: Intra-abdominal sepsis resulted in markedly less new collagen synthesis 1 day postanastomosis (9,163 +/- 1,234 versus 3,744 +/- 444 disintegrates per minute 3H-proline/mg of protein, P < 0.0001) and 4 days postanastomosis (8,462 +/- 956 versus 5,708 +/- 802 dpm/mg of protein P < 0.05). Noncollagenous protein synthesis was also impaired in anastomotic tissue from CLP rats on postanastomosis day 1 (37,497 +/- 3,740 versus 18,593 +/- 2,695 dpm/mg of of protein, P < 0.001) and postanastomosis day 4 (28,238 +/- 834 versus 17,784 +/- 1,415 dpm/mg of of protein, P < 0.0001). The expression of type I and type III collagen was altered relative to the normal temporal sequence observed in SHAM animals. CONCLUSION: Intra-abdominal infection impairs colonic reparative collagen and protein synthesis. In addition, regulation of type I and type III collagen genes is altered by intra-abdominal sepsis, and the alteration likely contributes to impaired new collagen synthesis and decreased colonic mechanical strength.

Failure of intestinal amino acid absorptive mechanisms in sepsis.

BACKGROUND: Sepsis has been shown to impair the barrier function and metabolism of the intestine. This study was done to investigate the effect of sepsis on intestinal absorption of proline, leucine, glutamic acid, and aminoisobutyric acid. STUDY DESIGN: Rats (six per group) were studied 24 hours after cecal ligation and puncture (CLP) or six hours after intraperitoneal injection of lipopolysaccharide (LPS). Controls underwent sham laparotomy or saline solution injection. Four 7-cm everted proximal jejunal sacs were prepared from each rat and filled with 800 microL Krebs' bicarbonate buffer containing 100 mumol/L of amino acid. Paired sacs (septic and control) were incubated at 37 degrees C in flasks containing the same solution trace labeled with 3H containing the same solution trace labeled with 3H amino acid. Sac contents were aspirated 60 minutes later and amino acid uptake was determined by scintillation counting. RESULTS: Twenty-four hours after CLP and six hours after LPS administration there was significant impairment in the intestinal absorption of all amino acids studied. Absorption of glutamic acid was the least affected, followed by leucine, aminoisobutyric acid, and proline. CONCLUSIONS: Sepsis impairs the intestinal absorption of amino acids. The magnitude of this defect in absorption differed with the amino acid studied, suggesting that not all transport systems were affected equally. This differential response of transport systems to sepsis appears to be the inverse of what is observed after a period of starvation.

Hypercalcemia decreases bile flow and increases biliary calcium in the prairie dog.

BACKGROUND: Biliary calcium is known to play an important role in the pathogenesis of gallstones. Calcium salts are present in all pigment gallstones and are also present in the core of most, if not all, cholesterol gallstones. METHODS: The effects of acute hypercalcemia on bile flow and biliary calcium secretion were examined in 22 prairie dogs during intravenous taurocholate infusion (0, 1.0, 2.25, and 4.5 mumol/kg/min). RESULTS: Bile flow was linearly correlated with bile acid output in both control (y = 7.62x + 13.5, r = 0.98) and hypercalcemic (y = 7.00x + 10.4, r = 0.96) animals. At lower bile acid outputs (< 3.0 mumol/kg/min), biliary ionized calcium output per increment bile acid output was significantly increased in hypercalcemic animals (0.016 versus 0.011 mumol Ca++ mumol taurocholate, p < 0.001). Bile ionized calcium concentrations approximated Gibbs-Donnan predicted values only at low bile flow rate. CONCLUSIONS: Hypercalcemia decreases bile flow and increases biliary ionized calcium concentration in the prairie dog. These effects favor the precipitation of calcium salts in bile.

Loss of colonic structural collagen impairs healing during intra-abdominal sepsis.

OBJECTIVE: To investigate colon anastomotic healing in an experimental model of sepsis. DESIGN: Prospective, randomized, experimental trial. SETTING: Experimental surgical laboratory of a large community hospital. STUDY PARTICIPANTS: Twenty-eight male Sprague-Dawley rats weighing 310 to 380 g. INTERVENTIONS: On day 0, the rats underwent either sham laparotomy or cecal ligation and puncture. The next day, the rats underwent left colon resection and single-layer inverted anastomosis. Colon-bursting pressure was determined 5 days after surgery at which time the anastomosis and a segment of colon 3 cm proximal to the anastomosis were excised. MAIN OUTCOME MEASURES: Colon-bursting pressure, colonic hydroxyproline concentration (index of collagen content), and total protein concentration measured as alpha-amino nitrogen. RESULTS: Sepsis resulted in decreased anastomotic bursting pressure and collagen concentration in all colon segments that were analyzed in the animals that underwent cecal ligation and puncture compared with control animals. CONCLUSIONS: Sepsis impairs healing of the colon, reflected by decreased bursting pressure and collagen concentration. The decrease in bowel wall structural collagen may affect the ability of the gut to hold sutures and thus may lead to more anastomotic failure.

Surgery--still an "old boys' club"?

BACKGROUND: Currently more than 40% of first-year medical students are female, yet less than 20% of those entering surgical residencies are women. It has been suggested that the surgical environment experienced during medical school clinical clerkships may be perceived as being unfavorable to female students, thus accounting for this disparity. METHODS: One hundred and five medical students at The Johns Hopkins University School of Medicine responded to a questionnaire administered before graduation to assess career choice, the influence of a number of conditions on choice of specialty, the perception of a specialty's attitudes toward their gender, and the students' experiences during clinical rotations. RESULTS: Thirty-four percent of men and 13% of women chose surgery or one of its subspecialties as their career (p < 0.01). Eighty-seven percent of women responding perceived a specialty as unfavorable toward their gender versus 21% of men (p < 0.0001). None of the men believed that surgery was unfavorable toward their gender, whereas 96% of these women believed that surgery was unfavorable (p < 0.00001). Fifty percent of women felt out of place on a clinical service, with 92% of these having this perception on a surgical service. Only 9% of men ever felt out of place on a clinical service (p < 0.0001), and none of the men felt out of place in surgery (p < 0.0001). CONCLUSIONS: Female medical students perceive a gender bias on surgical services, suggesting that an "old boys' club" attitude may still exist in surgery.

Effect of octreotide on sphincter of Oddi and gallbladder motility in prairie dogs.

Somatostatin and its synthetic analogue, octreotide, inhibit gallbladder emptying and cause gallstones. Whether octreotide-induced alterations in sphincter of Oddi motility contribute to this process is unknown. We, therefore, examined the effect of octreotide on fasting and protein-stimulated sphincter of Oddi motility. In 25 anesthetized prairie dogs, sphincter of Oddi motility and gallbladder pressure were monitored during the intravenous administration of octreotide, cholecystokinin (CCK) octapeptide, atropine, the intraduodenal administration of casein, and combinations of these agents. Intravenous octreotide decreased fasting sphincter of Oddi motility index both with (59 +/- 19 vs. 84 +/- 28, P less than 0.05) and without (137 +/- 31 vs. 227 +/- 42, P less than 0.05) prior cholinergic blockade with atropine. Octreotide also prevented the increases in sphincter of Oddi motility and gallbladder pressure seen with intraduodenal casein. Exogenous CCK increased sphincter of Oddi motility index and gallbladder pressure despite the simultaneous administration of octreotide alone (357 +/- 109 vs. 137 +/- 31, P less than 0.07, and 11.2 +/- 1.0 mmHg vs. 9.6 +/- 0.6 mmHg, P less than 0.05) or the combination of octreotide and atropine (317 +/- 69 vs. 59 +/- 19, P less than 0.05, and 10.1 +/- 1.6 mmHg vs. 8.5 +/- 1.4 mmHg, P less than 0.05). We conclude that both a cholinergic and an octreotide-sensitive noncholinergic pathway stimulate fasting sphincter of Oddi motility in the prairie dog.