Identifying novel therapeutic targets in gastric cancer using genome-wide CRISPR-Cas9 screening.

Genome-scale CRISPR-Cas9 screening technology is a powerful tool to systematically identify genes essential for cancer cell survival. Herein, TKOv3, a genome-scale CRISPR-Cas9 knock-out library, was screened in the gastric cancer (GC) cells, and relevant validation experiments were performed. We obtained 854 essential genes for the AGS cell line, and 184 were novel essential genes. After knocking down essential genes: SPC25, DHX37, ABCE1, SNRPB, TOP3A, RUVBL1, CIT, TACC3 and MTBP, cell viability and proliferation were significantly decreased. Then, we analysed the detected essential genes at different time points and proved more characteristic genes might appear with the extension of selection. After progressive selection using a series of open datasets, 41 essential genes were identified as potential drug targets. Among them, methyltransferase 1 (METTL1) was over expressed in GC tissues. High METTL1 expression was associated with poor prognosis among 3 of 6 GC cohorts. Furthermore, GC cells growth was significantly inhibited after the down-regulation of METTL1 in vitro and in vivo. Function analysis revealed that METTL1 might play a role in the cell cycle through AKT/STAT3 pathways. In conclusion, compared with existing genome-scale screenings, we obtained 184 novel essential genes. Among them, METTL1 was validated as a potential therapeutic target of GC.

Role of Tenascin-X in regulating TGF-ß/Smad signaling pathway in pathogenesis of slow transit constipation.

BACKGROUND: Chronic constipation is a gastrointestinal functional disease that seriously harms physical and mental health and impacts the quality of life of patients. Its incidence rate is 2%-27%. Slow transit constipation (STC) is a common type of chronic functional constipation, accounting for 10.3%-45.5% of such cases. Scholars have performed many studies on the pathogenesis of STC. These studies have indicated that the occurrence of STC may be related to multiple factors, such as dysfunction of the enteric nervous system, interstitial cells of Cajal (ICC) damage, and changes in neurotransmitters regulating intestinal peristalsis. AIM: To investigate the role of Tenascin-X (TNX) in regulating the TGF-ß/Smad signaling pathway in the pathogenesis of STC. METHODS: This study included an experimental group and a control group. The experimental group included 28 patients with severe colonic STC, and the control group included 18 patients with normal colon tissues. Immunohistochemistry (IHC) was used to detect c-Kit, a specific marker of the ICC. Western blot, immunofluorescence, and IHC were used to detect the localization and expression of TNX and TGF-ß/Smad. RESULTS: IHC showed that the number of ICC with positive c-Kit expression was significantly reduced in the colon of STC patients (22.17 ± 3.28 vs 28.69 ± 3.53, P < 0.05) and that the distribution was abnormal. Western blot results showed that c-Kit and Smad7 levels were significantly decreased in the colon of STC patients (c-kit: 0.462 ± 0.099 vs 0.783 ± 0.178, P < 0.01; Smad7: 0.626 ± 0.058 vs 0.799 ± 0.03, P < 0.01) and that TNX and Smad2/3 levels were higher in the STC group (TNX: 0.868 ± 0.028 vs 0.482 ± 0.032, P < 0.01). There was no significant difference in TGF-ß between the two groups (0.476 ± 0.028 vs 0.511 ± 0.044, P = 0.272). Pearson correlation analysis showed that the TNX protein exhibited a strong correlation with Smad2/3 and Smad7 (P < 0.05, |R| > 0.8) and TGF-ß (P < 0.05, |R| = 0.7). CONCLUSION: The extracellular matrix protein TNX may activate the TGF-ß/Smad signaling pathway by upregulating the Smad 2/3 signaling protein and thereby induce slight or complete epithelial stromal cell transformation, leading to an abnormal distribution and dysfunction of ICC in the diseased colon, which promotes the occurrence and development of STC.

Surgical outcomes of subtotal colectomy with antiperistaltic caecorectal anastomosis vs total colectomy with ileorectal anastomosis for intractable slow-transit constipation.

BACKGROUND: Few studies have compared the surgical outcomes of different surgical procedures currently used to treat refractory colonic slow-transit constipation (STC), despite the increase in the number of cases. This study aimed to analyse the long-term surgical outcomes of subtotal colectomy with antiperistaltic caecorectal anastomosis (SC-ACRA) vs total colectomy with ileorectal anastomosis (TC-IRA) for severe STC. METHODS: Between January 2005 and January 2015, we retrospectively collected clinical data of 55 patients who underwent TC-IRA (n = 35) or SC-ACRA (n = 20) for severe STC at our institution. The post-operative functional outcomes between the two groups were compared. RESULTS: There were no significant differences in age (P = 0.655), sex (P = 0.234), period of constipation (P = 0.105) and defecation frequency (P = 0.698) between the TC-IRA and SC-ACRA groups. During a median follow-up period of 72 months (range, 12-120 months), there were no significant differences between the TC-IRA and SC-ACRA groups regarding the median number of bowel movements per day [3 (1/6-7) vs 3 (1/6-5), P = 0.578], Cleveland Clinic Florida Constipation Score [2 (0-20) vs 2 (0-19), P = 0.454], Cleveland Clinic Incontinence Score [0 (0-5) vs 0 (0-2), P = 0.333] and Gastrointestinal Quality of Life Index [122 (81-132) vs 120 (80-132), P = 0.661]. Moreover, there was no significant difference in the incidence of post-operative complications between the two groups (37.1% vs 25.0%, P = 0.285). CONCLUSIONS: Our findings indicate that both TC-IRA and SC-ACRA are effective treatments for severe STC, with similar long-term outcomes.

Report of a rare case and review of adult intestinal duplication at the opposite side of mesenteric margin

ABSTRACT CONTEXT: To study the previously discovered clinical entity of adult intestinal duplication and its treatment, and propose an extension to its existing classification. CASE REPORT: We report the case of an adult male with abdominal pain, constipation and vomiting. This patient underwent surgical separation of adhesions, reduction of torsion and intestinal decompression. Postoperative pathological findings confirmed the rare diagnosis of intestinal duplication. CONCLUSION: Adult intestinal duplication is quite rare. Its clinical manifestations are nonspecific. From this finding of intestinal duplication originating at the opposite side of the mesenteric margin, a further extension of the existing anatomical classification is proposed.

Report of a rare case and review of adult intestinal duplication at the opposite side of mesenteric margin.

CONTEXT: To study the previously discovered clinical entity of adult intestinal duplication and its treatment, and propose an extension to its existing classification. CASE REPORT: We report the case of an adult male with abdominal pain, constipation and vomiting. This patient underwent surgical separation of adhesions, reduction of torsion and intestinal decompression. Postoperative pathological findings confirmed the rare diagnosis of intestinal duplication. CONCLUSION: Adult intestinal duplication is quite rare. Its clinical manifestations are nonspecific. From this finding of intestinal duplication originating at the opposite side of the mesenteric margin, a further extension of the existing anatomical classification is proposed.

Transanal surgery for obstructed defecation syndrome: Literature review and a single-center experience.

Obstructed defecation syndrome (ODS) is a functional disorder commonly encountered by colorectal surgeons and gastroenterologists, and greatly affects the quality of life of patients from both societal and psychological aspects. The underlying anatomical and pathophysiological changes of ODS are complex. However, intra-rectal intussusception and rectocele are frequently found in patients with ODS and both are thought to play an important role in the pathogenesis of ODS. With the development of evaluation methods in anorectal physiology laboratories and radiology studies, a great variety of new operative procedures, especially transanal procedures, have been invented to treat ODS. However, no procedure has been proved to be superior to others at present. Each operation has its own merits and defects. Thus, choosing appropriate transanal surgical procedures for the treatment of ODS remains a challenge for all surgeons. This review provides an introduction of the current problems and options for treatment of ODS and a detailed summary of the essential assessments needed for patient evaluation before carrying out transanal surgery. Besides, an overview of the benefits and problems of current transanal surgical procedures for treatment of ODS is summarized in this review. A report of clinical experience of some transanal surgical techniques used in the authors' center is also presented.

Correlation of Musashi-1, Lgr5, and pEGFR expressions in human small intestinal adenocarcinomas.

Recent studies have revealed that Musashi-1 and Lgr5 (leucine-rich-repeat-containing G-protein-coupled receptor 5) were putative stem cell genes. The epidermal growth factor receptor (EGFR) has also been extensively studied; it was known as an oncogenic driver in cancers. Overexpressions of Musashi-1, EGFR, and Lgr5 have been reported in some tumor tissues and cell lines. In this study, we used immunohistochemical analysis to investigate the expression pattern of Musashi-1, Lgr5, and pEGFR in 38 small intestinal adenocarcinomas (SIAs) resection specimens, 20 matched normal specimens and tried to analyze the correlations among them. The positive rate of Musashi-1, Lgr5, and pEGFR in SIAs, respectively, was 71 % (27/38), 55 % (21/38), and 45 % (17/38). Compared with the adjacent normal small intestinal mucosa, Musashi-1, Lgr5, and pEGFR protein were overexpressed in SIAs (P< 0.05). Furthermore, Musashi-1 and Lgr5 expressions were significantly correlated with the depth of wall invasion (P = 0.0011, P = 0.0017, respectively). Musashi-1 expression was closely correlated with Lgr5 (P = 0.015, r = 0.392). However, pEGFR expression was not associated with age, gender, tumor size, differentiation, depth of invasion, lymphatic metastasis, TNM stage, and pEGFR expression was not correlated with Musashi-1 or Lgr5 (P > 0.05, r = 0.064; P > 0.05, r = 0.307, respectively). Thus, we suggest that Musashi-1, Lgr5, and pEGFR are overexpressed in human SIAs and may play roles in human SIA carcinogenesis and progression.

[Total colonic exclusion plus side to side ileorectal antiperistaltic anastomosis in the treatment for elderly patients with slow transit constipation].

OBJECTIVE: To evaluate the feasibility and safety of total colonic exclusion plus side to side antiperistaltic ileorectal anastomosis in the treatment for elderly patients with slow transit constipation (STC). METHODS: Clinical data of 13 patients with severe idiopathic STC undergoing the above novel procedure in Zhongnan Hospital of Wuhan University between May 2009 and September 2012 were retrospectively analyzed. The Wexner constipation score and gastrointestinal quality of life index (GIQLI) before and 6 months after operation were compared. RESULTS: There were 8 female and 5 male patients, with a mean age of 74 years (range 63-82 years). No procedure-related deaths or any serious complication occurred. The length of follow-up ranged from 6 to 29 months (median,12 months). The duration of surgery was (55±4) min. Blood loss was (30±2) ml. The postoperative hospital stay ranged 10 to 16 days (mean 11.4 days). The first bowel movement occurred in the 4th day (range 2nd-8th day) after operation. There was no intestinal occlusion and anastomotic leakage that required surgery in all the patients. No fecal incontinence or constipation recurrence was found. One patient developed blind loop syndrome 14 months after operation. Postoperative complications included incision fat liquefaction in 2 cases, anorectal bearing-down while bowel movement in 2 cases, and minor defecate difficulty needed glycerin enema in 1 case. Wexner scores was significantly improved from 22.8±3.3 before operation to 5.4±2.1 six months after operation (P<0.05). GQLI was significantly increased from 93.6±20.5 before operation to 120.8±13.0 six months after operation (P<0.05). At 6 months after operation, the outcome was excellent in 11 patients and good in 2 patients. CONCLUSION: Total colonic exclusion plus side to side antiperistaltic ileorectal anastomosis is easy, safe and effective in the treatment for selected elderly patients with STC.

Molecular alterations of EGFR in small intestinal adenocarcinoma.

BACKGROUND AND AIMS: Molecular testing for epidermal growth factor receptor (EGFR) mutations has recently become a standard practice for the management of patients with non-squamous none small cell lung cancer. Primary small intestine adenocarcinoma (SIA) is an uncommon malignancy, and EGFR mutation in the cancer has not been well characterized due to its rarity. METHODS: A micro-tissue array with 53 SIAs and 24 surgically resected primary non-ampullary SIAs were studied. EGFR mutations were analyzed by DNA sequencing in 24 cases with formalin-fixed paraffin-embedded blocks. All 77 cases were examined by immunohistochemistry (IHC) using antibodies specific for the EGFR E746-A750 deletion in exon 19 (DEL), L858R point mutation in exon 21 (L858R), and total EGFR. EGFR amplifications were detected by fluorescence in situ hybridization. RESULTS: A positive reaction of DEL-specific, L858R-specific, and total EGFR antibodies was detected in seven (9.1%), 5 (6.5%) and 35 (45.5%) of 77 SIAs by IHC, respectively. Positive reaction of the three antibodies was not significantly correlated with patient's age, gender, differentiation, and stage. EGFR gene amplification was assayed in 77 SIAs in micro-tissue array. Of 24 SIA samples that had DNA sequencing, two (8.3%) harbored exon 19 deletion and one (4.2%) harbored L858R point mutation. Only one case with EGFR amplification and two cases with polysomy were shown. CONCLUSIONS: Our findings suggested that mutations and amplification in EGFR genes are minor events, and most of SIAs may be unsuitable to EGFR-TKIs treatment.

Current role of surgery for the treatment of fulminant Clostridium difficile colitis.

OBJECTIVE: This review discusses the current status and progress in studies on fulminant Clostridium difficile colitis (FCDC), including the definition, risk factor, diagnostic role of CT, surgical treatment, postoperative mortality, and new therapeutic strategy. DATA SOURCES: A literature search was conducted mainly in Medline and PubMed published in English between January 2000 and May 2011. The search terms were "ulminant Clostridium difficile colitis" "reatment", "urgery" and "ortality" RESULTS: Recent studies show that the overall mortality rate for FCDC remains high despite early surgical intervention. It has been difficult to identify the real value for surgical intervention in patients with FCDC due to the absence of prospective, randomized studies. Early recognition of patients with FCDC will help a clinician decide the need for treatment in an intensive care setting, multi-disciplinary consultation, and appropriate therapeutic selection. Some studies emphasize the importance of early recognition and emergent surgery at a less severe stage. Monoclonal antibody therapy and intravenous immunoglobulin treatment may be useful for the treatment of FCDC. CONCLUSIONS: Present studies do not provide strong evidence for guiding the surgical treatment of FCDC; hence, creation of collaborative research networks is crucial in order to undertake large prospective multi-center studies for improvement in overall survival.

[Experimental study of acellular dermal matrix biological patch enriched with bone marrow mesenchymal cells for the treatment of external anal sphincter injury].

OBJECTIVE: To investigate the feasibility of bone marrow mesenchymal stem cells(MSC)with the acellular dermal matrix(ADM) biological patch for the treatment of external anal sphincter injury on the animal models. METHODS: Thirty Wistar rats with sphincter injury were randomly divided into three groups. Group A underwent end to end sphincteric repair directly, group B underwent end to end repair and then covered by ADM patch, and group C underwent end to end repair and then covered by ADM which was previously seeded with MSC. After six weeks, the whole ring specimens including anal canal and lower rectum were removed. The hematoxylin and eosin stain and Masson trichrome stain were performed to observe the change of histomorphology. RESULTS: Two weeks later, the majority of rat models presented with moist anus and crissum with loose stools, which indicated that the model was established successfully. Six weeks after repair, in group A and B, the suffusion of fibrous connective tissue and the infiltration of inflammatory cells were observed at the repair site of sphincter. And lots of collagen fiber which was stained into blue deposited dispersedly at the site of repair with no obvious proliferation of capillaries. However, in group C, the blue collagenous fiber which deposited at the sphincter injury site was less than that in groups A and B. Muscle fibers were observed to be stained into red distributed dispersedly at the repair site of sphincter in group C. CONCLUSIONS: Transplantation of ADM biological patch rich in bone MSC can partly improve the regeneration of rat injured anal sphincter and lessen the formation of cicatrix.

[Use of antiperistaltic cecoproctostomy in colorectal reconstruction].

OBJECTIVE: To explore the feasibility and functional outcome of antiperistaltic cecoproctostomy in colorectal reconstruction. METHODS: Fifty-six patients who underwent antiperistaltic cecoproctostomy were retrospectively studied. Indications for antiperistaltic cecoproctostomy included slow transit constipation(n=44), synchronous colon cancer or colonic polyps(n=5), acute obstructing left colon carcinoma(n=4), and adult megacolon(n=3). RESULTS: Short-term postoperative complications included wound infections(n=5), 3 lymphatic leakages(n=3), and inflammatory small bowel obstruction(n=1). One month after antiperistaltic cecoproctostomy, the median frequency of daily bowel movement was 4.0(range, 2-6). After a median follow-up of 4 years(range, 1 month to 7 years), the median daily bowel frequency was 2.5(range, 0.5-4.0). Five patients suffered from long-term postoperative complications including small bowel obstruction(n=3), incision hernia(n=1), and mild cecal dilatation(n=1). The mean Wexner incontinence score was 4.2±1.1. CONCLUSION: Antiperistaltic cecoproctostomy is safe and effective for colorectal reconstruction.

[Association of vascular endothelial growth factor 936C/T polymorphism and the susceptibility to colorectal adenoma].

OBJECTIVE: To examine the association between polymorphism of vascular endothelial growth factor(VEGF)1498 C/T,936 C/T and colorectal adenoma genetic susceptibility. METHODS: A case-control study of 224 colorectal adenomas and 200 controls was conducted and VEGF genotypes were determined based on TaqMan-probe assay. The epidemiological factors were collected through questionnaire. Accordingly, the clinicopathological data of each sample were also investigated. RESULTS: The carriage of 936 CT and CT+TT genotypes had significantly higher risk of colorectal adenoma (CT vs. CC, OR=2.00, 95% CI: 1.23-3.25, P=0.006; CT+TT vs. CC, OR=2.04, 95% CI:1.28-3.26, P=0.003). 936-T allele carriage had increased risk of colorectal adenoma (OR=1.91, 95% CI:1.25-2.91, P=0.003). The genotypes of 1498 C/T and the frequency of C/T allele showed no differences between healthy persons and patients (P>0.05). In patients with 936 CT+TT and 936-T allele implied a tendency of villous adenoma category (CT+TT vs. CC, OR=2.54, 95% CI:1.12-5.75, P=0.040; T allele vs. C allele, OR=3.08, 95% CI, 1.64-5.80, P=0.001). CONCLUSION: VEGF 936 C/T polymorphism can influence susceptibility to colorectal adenoma.

Expression of putative stem cell genes Musashi-1 and beta1-integrin in human colorectal adenomas and adenocarcinomas.

BACKGROUND AND AIMS: Recent studies revealed that Musashi-1and beta1-integrin were putative stem cell genes. Overexpressions of Musashi-1 and beta1-integrin have been reported in some tumor tissues and cell lines. This study was to detect expressions of the two genes in colorectal adenomas and carcinomas and to analyze the correlation between Musashi-1 and beta1-integrin. METHODS: Musashi-1 and beta1-integrin immunoreactivity was studied immunohistochemically in tissue microarray-based samples containing 69 colorectal adenocarcinomas, eight normal mucosa, and eight adenomas, and their messenger RNA (mRNA) expression level was detected by RT-PCR in resected specimens including the three types of tissue. RESULTS: A percentage of 66.7% (46/69) and 59.2% (41/69) of colorectal adenocarcinomas were immunoreactive with Musashi-1 and beta1-integrin, respectively. The expressions of Musashi-1 and beta1-integrin protein were significantly higher in tissue samples of stage III than those of stage I-II (P = 0.0252; P = 0.0018, respectively). beta1-integrin expression was higher in group of adenocarcinomas than that of adenomas (P = 0.0276). Musashi-1 expression was closely correlated with beta1-integrin (rs = 0.631, P = 0.0001). Significant differences of Musashi-1 and beta1-integrin mRNA expression levels were found between the normal colorectal mucosa, adenoma, and adenocarcinoma tissues (P = 0.01; P = 0.03, respectively). CONCLUSIONS: Musashi-1 and beta1-integrin may be involved in human colorectal tumor carcinogenesis and progression. Our observations also indicate the need for further investigations to test in vivo whether cells with these markers have stem cell properties.

[Comparison of subtotal colectomy with antiperistaltic cecoproctostomy and total colectomy with ileoproctostomy in treating slow transit constipation].

OBJECTIVE: To compare clinical outcome and quality of life of subtotal colectomy with antiperistaltic cecoproctostomy and total colectomy with ileorectal anastomosis (TAC-IRA) in patients with severe slow transit constipation (STC). METHODS: Of the 56 patients enrolled in this study from January 1999 to June 2008, 32 cases underwent subtotal colectomy with antiperistaltic cecoproctostomy, and 20 patients underwent TAC-IRA. The patients' clinical characteristics, operative data, postoperative outcome, functional result and gastrointestinal quality of life index (GIQLI) survey were compared between the two groups. RESULTS: All patients were followed up for 1-7 years (median, 4 years). The basic clinical characteristics between the two groups was comparable. During the follow-up period, the number of daily bowel movements in the subtotal colectomy group was significantly fewer than that in TAC-IRA group (2.5+/-0.8 vs. 3.4+/-0.8; P=0.000). The Wexner continence score was significantly lower in subtotal colectomy group (4.4+/-1.6 vs. 5.8+/-1.9; P=0.011), and the GIQLI score in subtotal colectomy group was significantly higher than that in the TAC-IRA group (120.7+/-7.5 vs. 111.1+/-12.0; P=0.005). CONCLUSION: Subtotal colectomy with antiperistaltic cecoproctostomy appeared to be the superior treatment than the TAC-IRA for selected patients with slow transit constipation for improved functional outcomes and quality of life.

[Efficacy comparison of subtotal colectomy with antiperistaltic cecoproctostomy and total colectomy with ileorectal anastomosis for patients with slow transit constipation].

OBJECTIVE: To compare the efficacy between subtotal colectomy with antiperistaltic cecoproctostomy and total colectomy with ileorectal anastomosis (TAC-IRA) for patients with severe refractory slow transit constipation(STC). METHODS: During 1999 to 2002, TAC-IRA was the preferred procedure for 20 STC patients in our department. From 2003 to 2005, 17 STC patients underwent subtotal colectomy plus antiperistaltic cecoproctostomy. Clinical data of the two groups were collected and compared retrospectively. RESULTS: There were no significant differences in basic preoperative clinical data between the two groups. During the follow-up period, the time of daily defecation in the antiperistaltic cecoproctostomy group was less than that of TAC-IRA group (2.4+/-0.9 vs 3.4+/-0.8, P=0.0014), meanwhile the Wexner continence score was significantly lower in the antiperistaltic cecoproctostomy group (4.3+/-1.8 vs 5.8+/-1.9, P=0.0223). Barium enema after subtotal colectomy showed that residual ascending colon and cecum presented a sign of "reservoir". CONCLUSION: Subtotal colectomy with antiperistaltic cecoproctostomy is a better method for appropriately selected patients with STC than TAC-IRA.

Subtotal colectomy with antiperistaltic cecoproctostomy for selected patients with slow transit constipation-from Chinese report.

BACKGROUND AND AIMS: Total abdominal colectomy with ileorectal anastomosis (TAC-IRA) is recommended widely for the patients with severe, refractory slow transit constipation (STC). Subtotal colectomy with end-to-end antiperistaltic cecorectal anastomosis (Sarli procedure), an alternative for STC, has been paid particular attention. The purpose of this study was to retrospectively compare alterations of clinical functions and qualities of life between TAC-IRA and Sarli procedure. METHODS: Seventeen patients with STC who underwent Sarli procedure and 20 patients with STC who underwent TAC-IRA were chosen for this study. Patient characteristics, operative data, postoperative data, alterations of clinical function, and quality of life were compared. The gastrointestinal quality of life index (GIQLI) survey was used to evaluate postoperative qualities of life. RESULTS: At the mean 4-year follow-up (range 2-6 years), the frequency of daily bowel movement in the Sarli group was significantly less than that in the TAC-IRA group (2.4 +/- 0.9 vs. 3.4 +/- 0.8; P = 0.0014), and the Wexner continence scores were significantly lower in the Sarli group compared to the TAC-IRA group (4.3 +/- 1.8 vs. 5.8 +/- 1.9; P = 0.0223). However, the GIQLI score in Sarli group was higher than the TAC-IRA group (119.8 +/- 7.5 vs. 111.1 +/- 12.0, P = 0.0455). Post subtotal colectomy barium enema showed a sign of "reservoir" at the residual ascending colon and cecum. CONCLUSIONS: Compared to the TAC-IRA, subtotal colectomy with end-to-end antiperistaltic cecoproctostomy for appropriately selected patients with STC resulted in relief of constipation, satisfactory functional outcome, and improved qualities of life.

Role of Hypoxia-inducible factor-1 alpha and Survivin in colorectal carcinoma progression.

BACKGROUND AND AIMS: Hypoxia-inducible factor-1 alpha (HIF-1 alpha) is the main active subunit of HIF-1 that promoted tumor cells survival and critical steps in tumor progression and aggressiveness. The authors aimed to investigate the role of HIF-1 alpha and Survivin in colorectal cancer (CRC) progression. MATERIALS AND METHODS: Plasmid expressing small interfering RNA (siRNA) against HIF-1 alpha was constructed and transfected into LS174T cells with Lipofectamine. The LS174T cells were incubated for 24 h under hypoxic condition. The inhibitory effects of siRNA on HIF-1 alpha gene was determined by semiquantitative reverse transcriptase polymerase chain reaction and Western blot. Expression of HIF-1 alpha and Survivin was investigated by immunohistochemistry in colorectal adenocarcinomas tissue microarrays. RESULTS: HIF-1 alpha and Survivin expressions were markedly downregulated after the siRNA expression vector against HIF-1 alpha was transfected into the LS174T cells. Of the eight adenoma lesions, one case (12.25%) and four cases (50%) were positive for HIF-1 alpha and Survivin, respectively. Of the 69 cases of CRCs, 46 cases (66.7%) and 39 cases (56.5%) were positive for HIF-1 alpha and Survivin, respectively. The positive rate of HIF-1 alpha protein in CRCs was significantly higher than that in colorectal adenoma lesions (P < 0.05). HIF-1 alpha protein expression was significantly higher in patients with stage III than in patients with stage I-II CRCs (P < 0.01). In addition, overexpression of HIF-1 alpha in higher stages of CRCs was found to correlate positively with Survivin levels (P < 0.001). CONCLUSIONS: Our data demonstrate that HIF-1 alpha and Survivin are mostly expressed in invasive CRCs. Inhibition of HIF-1 alpha may lead to exploration of its potential as a diagnostic tool and possibly a target for gene therapy for colorectal carcinoma.

Anatomical variations of the cystic duct: two case reports.

Anatomical variations of the cystic duct often occur and may be encountered during cholecystectomy. Knowledge of the variable anatomy of the cystic duct and cysticohepatic junction is important to avoid significant ductal injury in biliary surgery. Here, we present two unusual cases with an anomalous cystic duct, namely, low lateral insertion and narrow-winding of the cystic duct. The first case was a 64-year-old man with cholelithiasis and chronic cholecystitis. During surgery, the entrance of the cystic duct was misidentified as being short and leading into the right hepatic duct. Further exploration showed multiple calculi in the right and common hepatic ducts. Cholecystectomy was completed, followed by T-tube drainage of the common and right hepatic ducts. Postoperative T-tube cholangiography demonstrated that the two T tubes were respectively located in the cystic and common hepatic duct. Six weeks later, the retained stones in the distal choledochus were extracted by cholangioscopy through the sinus tract of the T-tube. The second case was a 41-year-old woman, in which, preoperative endoscopic retrograde cholangiopancreatography (ERCP) revealed a long cystic duct, with a narrow and curved-in lumen. The patient underwent open cholecystectomy. Both patients were cured. The authors propose that preoperative ERCP or magnetic resonance cholangiopancreatography (MRCP), and intraoperative cholangiography or cholangioscopy constitute a useful and safe procedure for determining anatomical variations of the cystic duct.