Total Gastrectomy for CDH-1 Mutation Carriers: An Institutional Experience.

BACKGROUND: Gastric cancer is a leading cause of cancer-related death across the world. A subset of gastric cancers demonstrates an inherited genetic predisposition. Individuals with germline mutations in the CDH1 gene incur a lifetime risk for diffuse gastric cancer and benefit from prophylactic gastrectomy. The results for this operative intervention remain relatively undescribed in the literature, despite guidelines supporting its use. METHODS: We present a single-institution series of patients with confirmed CDH1 mutations who underwent gastrectomy. We describe their presenting symptoms, preoperative screening, clinicopathologic features, and outcomes. Focal outcomes of interest are weight loss and postoperative morbidity. RESULTS: Between 2010 and 2018, ten patients with a confirmed CDH1 mutation underwent total gastrectomy with intestinal pouch reconstruction at our institution. Two patients had clinical gastric cancer at the time of their operation at 21 and 60 y of age. Eight patients had prophylactic gastrectomy. All prophylactic patients had undergone prior endoscopic screening without detection of cancer; however, three had occult gastric cancer on pathological examination. Median weight loss after gastrectomy was 10 kg at 6 mo and 11 kg at 1 y. Postoperative morbidity was limited to one anastomotic leak, one hematoma, and one case of pneumonia. All patients remain disease-free with median follow-up of 19 mo. CONCLUSIONS: Total gastrectomy for patients with a CDH1 mutation is a cancer-preventing operation for a high-risk population. For this series, jejunal pouch reconstruction was performed with encouragingly low postoperative morbidity, weight loss, and good subjective function.

Acute Care Surgery for Transplant Recipients: A National Survey of Surgeon Perspectives and Practices.

BACKGROUND: Transplant recipients are living longer than ever before, and occasionally require acute care surgery for nontransplant-related issues. We hypothesized that while both acute care surgeons (ACS) and transplant surgeons would feel comfortable operating on this unique patient population, both would believe transplant centers provide superior care. METHODS: To characterize surgeon perspectives, we conducted a national survey of ACS and transplant surgeons. Surgeon- and center-specific demographics were collected; surgeon preferences were compared using χ2, Fisher's exact, and Kruskal-Wallis tests. RESULTS: We obtained 230 responses from ACS and 204 from transplant surgeons. ACS and transplant surgeons believed care is better at transplant centers (78% and 100%), and transplant recipients requiring acute care surgery should be transferred to a transplant center (80.2% and 87.2%). ACS felt comfortable operating (97.5%) and performing laparoscopy (94.0%) on transplant recipients. ACS cited transplant medication use as the most important underlying cause of increased surgical complications for transplant recipients. Transplant surgeons felt it was their responsibility to perform acute care surgery on transplant recipients (67.3%), but less so if patient underwent transplant at a different institution (26.5%). Transplant surgeons cited poor transplanted organ resiliency as the most important underlying cause of increased surgical complications for transplant recipients. CONCLUSIONS: ACS and transplant surgeons feel comfortable performing laparoscopic and open acute care surgery on transplant recipients, and recommend treating transplant recipients at transplant centers, despite the lack of supportive evidence. Elucidating common goals allows surgeons to provide optimal care for this unique patient population.

Irrigation Versus Suction in Laparoscopic Appendectomy for Complicated Appendicitis: A Meta-analysis.

BACKGROUND: Acute appendicitis is currently one of the most common surgical emergencies. Intra-abdominal abscesses (IAA) are a fearsome complication, which may occur. Irrigation during the appendectomy is one of the factors suggested to affect the rates of IAA. We sought to investigate the evidence regarding the use of irrigation versus suction alone and the development of IAA after laparoscopic appendectomy for complicated appendicitis. METHODS: We searched PubMed, Scopus, Embase, Cochrane, and the Web of Science through November 10, 2017, according to Preferred Reporting Items for Systematic Review and Meta-Analyses guidelines. We extracted information of interest, including rates of postoperative (IAA), and performed meta-analysis using random-effects model using the RevMan software. RESULTS: We identified five eligible studies with 2511 patients in total. Use of irrigation overall did not demonstrate significant increase in IAA (odds ratio [OR] = 2.39, 95% confidence interval [CI; 0.49, 11.74], P = 0.28). For the adult subpopulation, the use of irrigation was associated with nonsignificant lower odds of IAAs (OR = 0.42, 95% CI [0.15, 1.16]), whereas in pediatric with nonsignificant higher risk (OR = 2.98, 95% CI [0.25, 35.34]). Performance of irrigation led to the addition of, on average, 7 min to the duration of the operation (mean difference = 7.16, 95% CI [3.23, 11.09], P < 0.001). Irrigation did not affect postoperative length of stay (mean difference = -0.80, 95% CI [-2.30, 0.69], P = 0.29). CONCLUSIONS: Performance of irrigation during laparoscopic appendectomy does not seem to prevent the development of IAA in neither adults nor pediatric patients.

Low-dose gemcitabine induces major histocompatibility complex class I-related chain A/B expression and enhances an antitumor innate immune response in pancreatic cancer.

We investigated the effect of gemcitabine (GEM), a key drug for pancreatic cancer treatment, on the expression of cell surface MICA/B in pancreatic cancer cells and resulting cytotoxicity of γδ T cells. We assessed the effect of GEM on the upregulation of cell surface MICA/B expression by flow cytometry, utilizing six pancreatic cancer cell lines. MICA and CD16 expressions from resected pancreatic cancer patient specimens, which received neoadjuvant chemotherapy (NAC) with GEM, were analyzed by immunohistochemistry. GEM could increase MICA/B expression on cell surface in pancreatic cancer cell lines (in 2 of 6 cell lines). This effect was most effectively at concentration not affecting cell growth of GEM (0.001 µM), because MICA/B negative population was appeared at concentration at cytostatic and cytotoxic effect to cell growth (0.1 and 10 µM). The cytotoxic activity of γδ T cells against PANC-1 was detected and functions through interactions between NKG2D and MICA/B. However, the enhancement of NKG2D-dependent cytotoxicity with increased MICA/B expression, by GEM treatment, was not observed. In addition, soluble MIC molecules were released from pancreatic cancer cell lines in culture supernatant with GEM treatment. Immunohistochemical staining demonstrated that MICA expression in tumor cells and CD16 positive cells surrounding tumors were significantly higher in the NAC group compared to that of the control group. There was a significant correlation between NAC and MICA expression, as well as NAC and CD16 positive cell expression. The present results indicate that low-dose GEM-induced MICA/B expression enhances innate immune function rather than cytotoxicity in pancreatic cancer. In addition, our result suggests that the inhibition of cleavage and release of MIC molecules from the tumor surface could potentially improve NKG2D-dependent cytotoxicity.

Inpatient survival after gastrectomy for gastric cancer in the 21st century.

BACKGROUND: Surgical treatment for gastric cancer has evolved substantially. To understand how changes in patient- and hospital-level factors are associated with outcomes over the last decade, we examined a nationally representative sample. METHODS: Retrospective cross-sectional discharge data from the 2001-2010 Nationwide Inpatient Sample were analyzed using cross tabulation and multivariable regression modeling. Patients with a primary diagnosis of gastric cancer undergoing gastrectomy as primary procedure were included. We examined relationships between patient- and hospital-level factors, surgery type, and outcomes including in-hospital mortality and length of stay (LOS). RESULTS: A total of 67,327 patients with gastric cancer undergoing gastrectomy nationwide with complete information were included. Compared with patients treated in 2001, patients in 2010 were younger, more likely admitted electively, treated in a teaching hospital, or at an urban center. There was no difference in the type of procedure performed over time. Factors associated with an increased risk of in-hospital mortality included older age, male gender, and nonelective admission (P<0.05). In multivariable analysis, patients undergoing gastrectomy in 2010 demonstrated 40% lower odds of in-hospital mortality (odds ratio, 0.60; P=0.008). Overall mean LOS was 13.9 d (standard error, 0.1) without change over time. Factors associated with longer LOS included procedure type, hospital location, nonelective admission, and comorbid disease (all P<0.05). CONCLUSIONS: The adjusted odds of in-hospital mortality among surgically treated patients with gastric cancer decreased >40% between 2001 and 2010. Further research is warranted to determine if these findings are due to better patient selection, regionalization of care, or improvement of in-hospital quality of care.

LARP7 is a potential tumor suppressor gene in gastric cancer.

We previously reported frequent truncating mutations of the RNA-binding protein gene, La ribonucleoprotein domain family, member-7 (LARP7) in gastric cancers (GCs) with frequent microsatellite instability. LARP7 negatively regulates positive transcription elongation factor-b (p-TEFb) by binding to and stabilizing 7sk RNA. p-TEFb has been linked to proliferation and de-differentiation in various tissues. Therefore, we reasoned that loss of LARP7 may contribute to gastric tumorigenesis. In this study, we evaluated LARP7 mRNA expression in 18 GCs, their corresponding non-neoplastic gastric tissues (N(GC)), and 18 normal gastric tissues from healthy individuals (N(N)). We also assessed the effects of transient small interfering (siRNA)-mediated LARP7 knockdown in immortalized non-neoplastic gastric epithelial cells. LARP7 mRNA was significantly decreased in GCs (median 2.5) relative to N(N)s (median 14.9, P<0.01) as well as relative to their corresponding N(GC)s (median 8.1, P<0.01). Transfection of an siRNA directed against LARP7 (anti-LARP7 siRNA) into non-neoplastic gastric epithelial cells decreased 7sk levels by 72% relative to a control siRNA (P<0.01). Furthermore, anti-LARP7 siRNA transfection increased cell proliferation by 23% (P<0.01) and cell migration by 22% (P<0.001) relative to control siRNA transfection. Taken together, these findings suggest that LARP7 downregulation occurs early during gastric tumorigenesis and may promote gastric tumorigenesis via p-TEFb dysregulation.

Novel candidate colorectal cancer biomarkers identified by methylation microarray-based scanning.

DNA hypermethylation is a common epigenetic abnormality in colorectal cancers (CRCs) and a promising class of CRC screening biomarkers. We conducted a genome-wide search for novel neoplasia-specific hypermethylation events in the colon. We applied methylation microarray analysis to identify loci hypermethylated in 17 primary CRCs relative to eight non-neoplastic colonic mucosae (NCs) from neoplasia-free subjects. These CRC-associated hypermethylation events were then individually evaluated for their ability to discriminate neoplastic from non-neoplastic cases, based on real-time quantitative methylation-specific PCR (qMSP) assays in 113 colonic tissues: 51 CRCs, nine adenomas, 19 NCs from CRC patients (CRC-NCs), and 34 NCs from neoplasia-free subjects (control NCs). A strict microarray data filtering identified 169 candidate CRC-associated hypermethylation events. Fourteen of these 169 loci were evaluated using qMSP assays. Ten of these 14 methylation events significantly distinguished CRCs from age-matched control NCs (P<0.05 by receiver operator characteristic curve analysis); methylation of visual system homeobox 2 (VSX2) achieved the highest discriminative accuracy (83.3% sensitivity and 92.3% specificity, P<1×10(-6)), followed by BEN domain containing 4 (BEND4), neuronal pentraxin I (NPTX1), ALX homeobox 3 (ALX3), miR-34b, glucagon-like peptide 1 receptor (GLP1R), BTG4, homer homolog 2 (HOMER2), zinc finger protein 583 (ZNF583), and gap junction protein, gamma 1 (GJC1). Adenomas were significantly discriminated from control NCs by hypermethylation of VSX2, BEND4, NPTX1, miR-34b, GLP1R, and HOMER2 (P<0.05). CRC-NCs were significantly distinguished from control NCs by methylation of ALX3 (P<1×10(-4)). In conclusion, systematic methylome-wide analysis has identified ten novel methylation events in neoplastic and non-neoplastic colonic mucosae from CRC patients. These potential biomarkers significantly discriminate CRC patients from controls. Thus, they merit further evaluation in stool- and circulating DNA-based CRC detection studies.

A novel sensor kinase is required for Bordetella bronchiseptica to colonize the lower respiratory tract.

Bacterial virulence is influenced by the activity of two-component regulator systems (TCSs), which consist of membrane-bound sensor kinases that allow bacteria to sense the external environment and cytoplasmic, DNA-binding response regulator proteins that control appropriate gene expression. Respiratory pathogens of the Bordetella genus require the well-studied TCS BvgAS to control the expression of many genes required for colonization of the mammalian respiratory tract. Here we describe the identification of a novel gene in Bordetella bronchiseptica, plrS, the product of which shares sequence homology to several NtrY-family sensor kinases and is required for B. bronchiseptica to colonize and persist in the lower, but not upper, respiratory tract in rats and mice. The plrS gene is located immediately 5' to and presumably cotranscribed with a gene encoding a putative response regulator, supporting the idea that PlrS and the product of the downstream gene may compose a TCS. Consistent with this hypothesis, the PlrS-dependent colonization phenotype requires a conserved histidine that serves as the site of autophosphorylation in other sensor kinases, and in strains lacking plrS, the production and/or cellular localization of several immune-recognized proteins is altered in comparison to that in the wild-type strain. Because plrS is required for colonization and persistence only in the lower respiratory tract, a site where innate and adaptive immune mechanisms actively target infectious agents, we hypothesize that its role may be to allow Bordetella to resist the host immune response.

Vaccine impedes the development of reflux-induced esophageal cancer in a surgical rat model: efficacy of the vaccine in a Pre-Barrett's esophagus setting.

BACKGROUND & AIMS: We developed a granulocyte-macrophage-colony-stimulating factor (GM-CSF) tumor vaccine for esophageal cancer. We evaluated the effectiveness of the vaccine as a prevention option in a surgical reflux rat model of esophageal cancer. METHODS: A surgical model involving a jejuno-esophagostomy was used to create Barrett's esophagus and esophageal cancer in rats. No carcinogen exposure was utilized. Cell lines derived from these tumors were stably passaged in vitro. GM-CSF-secreting tumor cells were generated using stable transfection. All rats underwent a total gastrectomy, followed by a jejuno-esophagostomy. The surgery promoted the reflux of duodenal contents into the esophagus. All animals were administered either a GM-CSF secreting whole cell vaccine or a phosphate-buffered saline (PBS) placebo injection 4, 6, 14, and 16 weeks post-surgery. RESULTS: While 15 of 16 animals in the non-vaccinated placebo group developed esophageal cancer, 94% (15 of 16), animals in the vaccine group had an incidence of cancer of 25% (4 of 16) (p<0.05). Barrett's esophagus was seen in 100% (16 of 16) of the controls and 83% (13 of 16) of the vaccinated animals. CONCLUSIONS: A GM-CSF-secreting whole cell tumor vaccine impeded esophageal tumor growth, but not the development of Barrett's esophagus, in a clinically relevant surgical reflux model.

Delivery of plasmid DNA expression vector for keratinocyte growth factor-1 using electroporation to improve cutaneous wound healing in a septic rat model.

We have previously shown that wound healing was improved in a diabetic mouse model of impaired wound healing following transfection with keratinocyte growth factor-1 (KGF-1) cDNA. We now extend these findings to the characterization of the effects of DNA plasmid vectors delivered to rats using electroporation (EP) in vivo in a sepsis-based model of impaired wound healing. To assess plasmid transfection and wound healing, gWIZ luciferase and PCDNA3.1/KGF-1 expression vectors were used, respectively. Cutaneous wounds were produced using an 8 mm-punch biopsy in Sprague-Dawley rats in which healing was impaired by cecal ligation-induced sepsis. We used National Institutes of Health image analysis software and histologic assessment to analyze wound closure and found that EP increased expression of gWIZ luciferase vector up to 53-fold compared with transfection without EP (p < 0.001). EP-assisted plasmid transfection was found to be localized to skin. Septic rats had a 4.7 times larger average wound area on day 9 compared with control (p < 0.001). Rats that underwent PCDNA3.1/KGF-1 transfection with EP had 60% smaller wounds on day 12 compared with vector without EP (p < 0.009). Quality of healing with KGF-1 vector plus EP scored 3.0 +/- 0.3 and was significantly better than that of 1.8 +/- 0.3 for treatment with vector alone (p < 0.05). We conclude that both the rate and quality of healing were improved with DNA plasmid expression vector for growth factor delivered with EP to septic rats.

The role of magnetic resonance cholangiography in the management of patients with gallstone pancreatitis.

OBJECTIVE: To examine the utility of magnetic resonance cholangiography (MRC) in the preoperative evaluation of patients with gallstone pancreatitis. SUMMARY BACKGROUND DATA: Gallstone pancreatitis is often associated with the presence of common bile duct (CBD) stones that may require endoscopic removal prior to planned laparoscopic cholecystectomy. No reliable clinical criteria exist, however, that can accurately predict CBD stones and the need for preoperative endoscopic retrograde cholangiopancreatography (ERCP). METHODS: Sixty-four patients were identified with gallstone pancreatitis based on clinical presentation and imaging studies over a three-and-a-half-year period. All patients underwent MRC, and the images were evaluated for gallstones, CBD stones, cholecystitis, and pancreatitis RESULTS: Seventeen of the 64 patients (27%) with gallstone pancreatitis were found to have CBD stones confirmed by ERCP. MRC correctly predicted CBD stones in 16 of the 17 patients (sensitivity = 94%). In 1 additional patient, MRC demonstrated CBD stones not seen at ERCP, consistent with probable passage. By comparison, the sensitivities of other criteria for predicting CBD stones were (1) elevated bilirubin >or=2.0 mg/dL = 65%; (2) dilated duct on ultrasound = 55%; and (3) CBD stones on ultrasound = 27%. MRC was able to visualize gallbladder stones in 57 of 62 patients (94%) and correctly predicted acute cholecystitis in 6 of 8 patients. MRC also detected peripancreatic edema and inflammatory changes consistent with acute pancreatitis in 45 of 64 patients (70%). CONCLUSIONS: These results demonstrate that MRC can accurately identify CBD stones preoperatively in patients with gallstone pancreatitis and provide valuable information with respect to other biliary pathology, including cholelithiasis, acute cholecystitis, and pancreatitis. MRC is an effective noninvasive screening tool for CBD stones, appropriately selecting candidates for preoperative ERCP and sparing others the need for an endoscopic procedure with its associated complications.

Electroporation enhances transfection efficiency in murine cutaneous wounds.

Transfection of wounds with DNA-encoding growth factors has the potential to improve healing, but current means of nonviral gene delivery are inefficient. Repeated high doses of DNA, necessary to achieve reliable gene expression, are detrimental to healing. We assessed the ability of in vivo electroporation to enhance gene expression. Full-thickness cutaneous excisional wounds were created on the dorsum of female mice. A luciferase- encoding plasmid driven by a CMV promoter was injected at the wound border. Following plasmid administration, electroporative pulses were applied to injection sites. Pulse parameters were varied over a range of voltage, duration, and number. Animals were euthanized at intervals after transfection and the luciferase activity measured. Application of electric pulses consistently increased luciferase expression. The electroporative effect was most marked at a plasmid dose of 50 micro g, where an approximate tenfold increase was seen. Six 100- micro s-duration pulses of 1750 V/cm were found to be the most effective in increasing luciferase activity. High numbers of pulses tended to be less effective than smaller numbers. This optimal electroporation regimen had no detrimental effect on wound healing. We conclude that electroporation increases the efficiency of transgene expression and may have a role in gene therapy to enhance wound healing.

Duodenal reflux produces hyperproliferative epithelial esophagitis--a possible precursor to esophageal adenocarcinoma in the rat.

Esophageal reflux of duodenal contents converts a rat nitrosamine esophageal cancer model from squamous cell carcinoma to adenocarcinoma. Further, there was a tendency for male rats to have a higher incidence of cancer than female rats. However, chemical castration with the gonadotropin-releasing hormone analog leuprolide did not protect male or female animals from developing cancer. We have identified an early (6-week) hyperproliferative epithelial cell reaction to duodenal reflux. We carried out experiments to assess the specificity of duodenal reflux in producing the hyperproliferative epithelial precursor lesion. Animals underwent specific surgical procedures to produce esophageal reflux of pure duodenal contents, mixed gastroduodenal, or bland intestinal contents. A hyperproliferative mucosal esophagitis developed in the group with duodenal reflux but not in the other groups. Mucosal thickness in the duodenal reflux group reached seven times that of normal mucosa at 6 weeks. These results suggest that esophageal reflux of duodenal contents plays an important role in the pathogenicity of proliferative esophagitis and the potential development of esophageal adenocarcinoma.

Increased laparoscopic experience does not lead to improved results with acute cholecystitis.

BACKGROUND: As laparoscopic experience increases, ever more challenging cases are attempted. Enlarged surgeon experience, along with better technology, has been lauded as improving outcomes. The purpose of this study is to see if this applies to the management of acute cholecystitis. METHODS: We reviewed our experience over a 7 and a half year period. Information was obtained from a prospectively maintained computer database containing patient presentation, demographics, workup, laboratory values, and outcomes. Diagnosis of gangrene was based on pathologic examination of the specimen. RESULTS: There were 305 patients admitted to our institution with acute cholecystitis. Group I (n = 111) was admitted during the first half of the study and group II (n = 194) during the second half. Demographics were similar in the two groups. While slightly more patients were attempted laparoscopically in group II (90% versus 82%), conversion rates were virtually identical (27.1% versus 27.5%). There was a trend toward improved results with group II versus group I in mortality (3% versus 4%) and morbidity (14% versus 21%; P = not significant). Deaths were divided between sepsis and cardiac events. Gangrenous cholecystitis was less frequent in group II patients (29% versus 40%; P = 0.06). Analysis of gangrene versus non-gangrene patients within each group showed that conversion rates remained twice as high (40% versus 20%; P < 0.05) in those with gangrene. Interestingly, gangrene had no effect on morbidity or mortality. CONCLUSIONS: Morbidity and mortality for acute cholecystitis remain relatively high. These seem to be determined by the degree of acute and chronic illness present at the time of diagnosis. As conversion rates remain unchanged, increased surgeon experience and further advances in laparoscopic technology are unlikely to dramatically affect results. Efforts to improve outcomes for this common disease should therefore focus on better and earlier identification of patients for operation.

A nuclear targeting peptide, M9, improves transfection efficiency in fibroblasts.

BACKGROUND: Nonviral transfection of eukaryotic cells remains inefficient. Liposomes can transport DNA plasmid into the cytoplasm, but the nuclear membrane remains a barrier to efficient plasmid DNA transfection. But normal cells have mechanisms to transport nucleic acids across the nuclear membrane. Cells routinely utilize a transporter to carry mRNA from nucleus to cytoplasm. MATERIALS AND METHODS: We used a modified mRNA transporter, the M9 component of heterogeneous nuclear ribonucleoprotein-A1, complexed to a DNA carrier to facilitate DNA transfer into the nucleus. We examined the effect of M9 on transfection in 3T3 fibroblasts. Our hypothesis was that the M9 shuttle would increase transfection efficiency by delivering plasmid to the nucleus, after cytoplasmic entry was facilitated by Lipofectamine. Transfection was assessed using plasmids expressing beta-galactosidase and green fluorescent protein (GFP). Intracellular location of rhodamine-labeled plasmid was determined by fluoroscopic microscopy. RESULTS: In the fluorescent microscopy experiments, we found that rhodamine-labeled DNA plasmid was sequestered in the cytoplasm in the Lipofectamine-treated cells, but gained access to the nucleus with the addition of M9. At concentrations where neither M9 nor Lipofectamine individually increased plasmid mediated transfection, as evidenced by beta-galactosidase activity; their combination increased transfection dramatically by approximately 20-fold, from 2 +/- 1 to 32 +/- 5. CONCLUSIONS: As expected, based on their presumed actions, Lipofectamine and the M9 shuttle synergistically promote efficient cellular transfection. Efficient cellular transfection will be required in clinical applications of gene therapy.