Emina H. Huang

Safety and reliability of tattooing colorectal neoplasms prior to laparoscopic resection

Accurate tumor localization is critical to performing minimally invasive colorectal resection. This study reviews the safety and reliability of tattooing colorectal neoplasms prior to laparoscopic resection. Weretrospectively reviewed 50 consecutive patients with colorectal neoplasms who underwent endoscopic tattooing prior to laparoscopic resection. Data were obtained from medical charts, endoscopy records, and pathology reports. No complications related to endoscopy or tattooing were incurred. Five neoplasms (10%) were in the ascending colon, five (10%) were in the transverse colon, eight (16%) were in the descending colon, 23 (46%) were in the sigmoid colon, and nine (18%) were in the rectum. Tattoos were visualized intraoperatively and accurately localized the neoplasm in 44 patients (88%). Six patients (12%) did not have tattoos visualized laparoscopically and required intraoperative localization. On average, the pathology specimens in this series had a 15 cmproximal margin, a 12 cmdistal margin, and 15 lymph nodes. In the context of laparoscopic colorectal resection, preoperative endoscopic tattooing is a safe and reliable method of tumor localization in most cases. Localizing colon and proximal rectal lesions with tattoos may be preferable to other localization techniques including intraoperative endoscopy.

Laparoscopic-assisted cecectomy is associated with decreased formation of postoperative pulmonary metastases compared with open cecectomy in a murine model

BACKGROUND It was shown in a murine model that sham laparotomy is associated with a higher incidence of postoperative lung metastases when compared with results seen after carbon dioxide pneumoperitoneum. Using the same tumor model, the present study was undertaken to determine if the addition of bowel resection to the operative procedure would impact the results. METHODS Sixty mice underwent anesthesia alone (anesthesia control [AC]), laparoscopic-assisted cecectomy (LC), or open cecectomy (OC). After surgery, all animals received tail vein injections of 105 TA3-Ha tumor cells. On postoperative day 14, the lungs and trachea were excised en bloc and processed, and surface lung metastases were counted and recorded by a blinded observer. RESULTS The mean number of surface lung metastases in the AC, LC, and OC groups was 30.9, 76.3, and 134.5, respectively. Significantly more metastases were documented after OC (P<.001) and LC (P<.05) than after anesthesia alone. Mice in the LC group had significantly fewer lung metastases (43% less) than mice in the OC group (P<.01). CONCLUSIONS OC was associated with significantly more lung metastases than either LC or AC. Surgery-related immune suppression or trophic tumor cell stimulation occurring after surgery may contribute to this phenomenon.

Colonoscopy in Mice

Introduction: Current investigational models of murine colitis and colon cancer necessitate sacrifice of animals in order to obtain colonic tissue. The purpose of this study was to develop a safe method of murine colonoscopy that would allow serial evaluation and mucosal biopsies of the same animal. Methods: Nine mice (two C3H, two C57/BL6, and five IL-10 deficient) were studied a total of four times each over 4 weeks. Three mice [APC (Min ±)] were examined three times each. Mice were gavaged with 1 cc of a polyethylene glycol solution on the day prior to colonoscopy. Solid chow was withheld and the mice were maintained on Pedialyte. Mice were anesthetized with ketamine and xylazine. A flexible pediatric cystoscope (2.1-mm diameter) with a single biopsy channel was introduced per anum, and the colon was gently insufflated with air to a mean pressure of less than 5 mmHg. Saline irrigation was used when necessary. A single biopsy was obtained from the rectosigmoid colon during each examination. Results: A total of 46 examinations were carried out. One mouse died after being anesthesized for the fourth examination, and two mice [one IL-10 knockout and one APC (Min±)] died one day after the 3rd examination. No other complications were noted. The average length of insertion was 3 cm. Transillumination allowed for localization of the endoscope tip. Biopsies, although quite small, were sufficient for pathologic evaluation and diagnosis. Conclusions: Murine colonoscopy is a safe and feasible technique. It permits consecutive visual and histopathological examinations, and it allows the investigator to monitor the response of the murine colon to experimental interventions.

Hybrid laparoscopic flexure takedown and open procedure for rectal resection is associated with significantly shorter length of stay than equivalent open resection

PURPOSE: Laparoscopic-assisted, sphincter-saving resection (largest incision <7 cm) of the middle and distal rectum is technically very difficult and, with regard to cancers, has not been demonstrated to be oncologically safe. The hypothesis of this retrospective study is that a hybrid operation that combines laparoscopic and open methods would be associated with short-term outcome benefits compared with open surgery results for patients undergoing sphincter-saving proctectomy. METHODS: A total of 31 hybrid and 25 fully open rectal resection patients were compared in this retrospective review. All patients had splenic flexure takedown and rectal anastomosis. The hybrid approach consisted of laparoscopic splenic flexure takedown (with or without partial rectal mobilization and devascularization) followed by completion of the procedurevia infraumbilical midline laparotomy. The indication was neoplasm in 87 percent of hybrid patients and in 68 percent of open patients. The majority of tumors were located between 4 and 10 cm from the dentate line. RESULTS: Fifty-eight percent of hybrid and 68 percent of open patients had low anterior or coloanal resections, and 48 percent of hybrid and 64 percent of open patients underwent temporary diversionvia ileostomy. The mean hybrid midline incision length was 11 cm compared with 24 cm for open patients (P <0.0001). The neoplastic specimens were similar with regard to margins and lymph node harvest. Similar complication rates were noted in both groups. Nonsignificant benefits for hybrid patients (0.9–1.2 days) were seen with regard to length of time until toleration of liquid or solid diet and first flatus. Hybrid patients experienced their first bowel movements 4.1 daysvs. 5.7 days for the open group (P=0.03). Mean length of stay was significantly shorter for hybrid patients (6.1. days) than for open patients (11.1 days;P=0.0006). CONCLUSION: This preliminary retrospective study suggests that a combined hybrid laparoscopic and open approach to sphincter-saving proctectomy permits a similar resection as open methods and may be associated with a length-of-stay benefit and more rapid return of bowel function. Prospective studies will be needed before any firm conclusions can be drawn.

The percentage of CD31 + T cells decreases after open but not laparoscopic surgery

Background: Efficient killing of tumor cells depends on T cells that migrate from the circulation to the peripheral tissues; these cells express CD31. This study was undertaken to determine the impact of open (OS) and laparoscopic (LS) colorectal surgery on the percentage of circulating CD3+CD31+ cells. Methods: Peripheral blood was collected from 27 OS and 24 LS colon cancer patients preoperatively (preOP) and on postoperative days 1 (POD1) and 3 (POD3). CD31+ T cells were assessed by flow cytometry using monoclonal antibodies. Results: In the OS group, the percentage of CD3+CD31+ cells was significantly lower in POD1 and POD3 samples compared to the preOP results. LS surgery did not result in a significant change in the percentage of these T cells. A significant correlation was found between the decrease in the percentage of CD3+CD31+ cells and the length of incision in OS patients. Conclusions: The percentage of CD3+CD31+ cells decreases following OS but not LS and may be related to incision length. This may compromise T cell function in the peripheral tissues in the postoperative period.

Increased platelet-derived growth factor (PDGF) release after laparotomy stimulates systemic tumor growth in mice

BackgroundOur laboratory has demonstrated that tumors grow larger and are more easily established following laparotomy than after carbon dioxide (CO2) pneumoperitoneum or anesthesia alone. We have also shown that tumor cells incubated with serum from laparotomized mice proliferated significantly faster in vitro than those incubated with plasma from mice that underwent laparoscopy or anesthesia alone. We hypothesized that differing levels of a plasma-soluble growth factor(s) postoperatively causes tumors to proliferate faster after laparotomy. This study’s purpose was to isolate and characterize the plasma growth factor(s) responsible for the increased growth of systemic tumors after laparotomy.MethodsFemale Balb/C mice (n=100) were randomized to two groups: anesthesia control (AC) or midline sham laparotomy (4 cm) (Open). Plasma was collected on Postoperative day 4. For the tumor proliferation assay, C-26 colon cancer cells were incubated in media with either 10% AC or Open “raw” plasma (not passed through column), or AC or Open plasma that had been passed through the column. For elution of heparin-binding proteins, plasma from each group was passed through a heparin-sepharose column. Elution of bound proteins was accomplished with a 0.1–2 M NaCl gradient. Each fraction was examined for protein content. For the anti-platelet-derived growth factor (PDGF) neutralizing antibody study, C-26 cells were incubated with one of four plasma preparations: AC or Open plasma alone, or AC or Open plasma incubated with anti-PDGF antibody. For both studies, tumor proliferation was determined after 2 days with an MTS/PMS assay. Results from each group were compared and differences determined using analysis of variance (ANOVA) and Tukey-Kramer tests.ResultsOn heparin chromatography, a single elution peak consistent with PDGF was present in both AC and Open plasma and was 1.5 times greater in the Open plasma. The first tumor proliferation assay showed that tumor cells incubated with Open plasma proliferated 2.5 times faster than those with AC plasma (p<0.0001). Passage of AC plasma through the column did not alter its mitogenic activity, but Open plasma thus treated demonstrated significantly decreased mitogenic activity. The second tumor proliferation assay showed that anti-PDGF antibody had no effect on the mitogenic activity of the AC plasma but decreased the mitogenic activity of the Open plasma to the AC plasma level.ConclusionsLaparotomy is associated with higher levels of a heparin-binding plasma factor, consistent with PDGF. The enhanced mitogenic activity of the OP plasma was neutralized with anti-PDGF antibody. Increased plasma levels of PDGF after laparotomy may be responsible for accelerated tumor growth following laparotomy in mice.

Perioperative Immunomodulation With Flt3 Kinase Ligand or a Whole Tumor Cell Vaccine Is Associated With a Reduction in Lung Metastasis Formation After Laparotomy in Mice

Introduction:Laparotomy has been associated with temporary postoperative immunosuppression and accelerated tumor growth in experimental models. In a previous murine study, a whole cell vaccine plus the adjuvant monophosphoryl-lipid A was shown to be effective in decreasing the number of lung metastases that develop after laparotomy. This study was conducted to assess the impact of the adjuvant fetal liver tyrosine kinase 3 (Flt3) ligand on perioperative tumor growth when used alone or with a tumor cell vaccine. Methods:An intravenous tumor cell injection lung metastases model was used. Sixty female A/J mice were divided into six equal groups designated (1) anesthesia control (AC), (2) AC with Flt3 ligand (ACFlt3), (3) sham laparotomy (OP), (4) OP with Flt3 ligand (OPFlt3), (5) OP with vaccine (OPVac), and (6) OP with Flt3 ligand and vaccine (OPFlt3Vac). Groups 2, 4, and 6 received daily intraperitoneal injections of Flt3 ligand (10μg/dose with carrier) for 5 days before and 5 days after surgery. Groups 1 and 3 received similar injections of saline on the same schedule. Groups 5 and 6 were vaccinated with irradiated whole Ta3Ha tumor cells intraperitoneally three times before and twice after surgery. Immediately after surgery, all mice were injected with 105 Ta3Ha tumor cells via a tail vein. After 14 days, the mice were sacrificed and their lungs and tracheas were excised en bloc. Specimens were stained and counterstained with India ink and Fekete solution, and surface metastases were counted by a blinded observer. Differences between study groups were determined by analysis of variance. The peritumoral inflammatory cell infiltrate of some Flt3 and control specimens was also assessed. Results:Regarding laparotomy, Flt3 ligand (mean, 1.22 metastases), whole cell vaccine (1.12 metastases), and the combination of these two agents (0.1 metastases) were each effective in significantly decreasing the number of surface lung metastases compared with surgery alone (9.88 metastases, P < .05 for all comparisons). There were no differences between the various treatment groups in regards to number of metastases. Only the combination of Flt3 and the vaccine significantly lowered the incidence of tumors (number of mice with>1 tumors). Histologic analysis revealed that the Flt3-treated mice demonstrated increased numbers of antigen-presenting cells surrounding the tumors compared with controls. Conclusions: Perioperative treatment with either Flt3 ligand or a whole cell tumor vaccine significantly reduced the number of lung metastases after laparotomy. The combination of the Flt3 ligand and the vaccine also decreased the incidence of metastases and was the most effective treatment. Further studies regarding perioperative immune modulation in the setting of cancer appear warranted.

The diagnostic and therapeutic roles of colonoscopy

Examination of the anus and rectum using various instruments has been available since the time of the Egyptians and Romans. Until the development of incandescent light, visualization was difficult, using only mirrors or candlelight. The advent of electricity allowed for development of a lighted rectosigmoidoscope by James P. Tuttle in 1903 [60]. Visualization proximal to the rectosigmoid junction, however, was difficult due to the rigidity of the instrument and the anatomy of the rectosigmoid junction and the sigmoid colon. In 1928, Hoff [35], a radiologist, used a rubber tube to intubate the cecum in a retrograde fashion [9] using fluoroscopic guidance. However, Bergein F. Overholt is credited with the development of a flexible, fiberoptic sigmoidoscope in 1963 [71].

Significant Reduction of Laparotomy-Associated Lung Metastases and Subcutaneous Tumors After Perioperative Immunomodulation with Flt3 Ligand in Mice

Laparotomy has been associated with increased rates of tumor establishment and metastasis formation postoperatively in animal models. The purpose of this study was to determine the impact on postoperative tumor growth of perioperative upregulation of immune function via fetal liver tyrosine kinase 3 (Flt3 ligand). Two murine studies were carried out: the first utilized a lung metastases model, and the second involved a subcutaneous tumor model. Each study included four groups: anesthesia control (AC), AC plus Flt3 ligand (ACFlt3), sham laparotomy (OP), and OP plus Flt3 ligand (OPFlt3). Flt3 ligand was administered by daily intraperitoneal injection (10 µg/dose) beginning 5 days preoperatively and continuing for 1 week postoperatively. In study 1, A/J mice were given tail vein injections of 1.5 x 105 TA3Ha cancer cells on the day of surgery. The mice were sacrificed 14 days after surgery, the lungs processed, and the surface metastases counted by a blinded observer. In study 2 C3H/He mice were given a dorsal subcutaneous injection of 104 MC-2 cancer cells on the day of surgery. The mice were sacrificed 31 days after surgery, and the injection sites were evaluated for subcutaneous tumors grossly and histologically. In study 1, the median number of surface lung metastases per mouse was 166 in the OP group and 38 in the OPFlt3 (P= .021). Mice in the AC group developed a median 50 lung metastases per animal compared with mice in the ACFlt3 group who had a median of 10 metastases per mouse (P= .001). The OP group had significantly more metastases than the AC group (P= .048). In study 2, the percentage of animals that developed tumors in the AC, OP, ACFlt3, and OPFlt3 groups was 43, 80, 0, and 20, respectively. The incidence of tumors in the OPFLt3 group and the ACFlt3 group was significantly less than their respective control groups (P< .01). The difference between the OP and AC groups was not significant (P> .05). Perioperatively administered Flt3 ligand was associated with significantly fewer lung metastases and a lower incidence of subcutaneous tumor formation after laparotomy and anesthesia alone. Perioperative immunomodulation may limit untoward surgery-related oncologic effects.