Impact of Preoperative Three-Dimensional Endoanal Ultrasound on the Surgical Outcome of Primary Fistula in Ano. A Multi-Center Observational Study of 253 Patients.

PURPOSE: To evaluate the impact of preoperative three-dimensional endoanal ultrasound (3D-EAUS) on the clinical outcome of anal fistula surgery. METHODS: The research consisted of multi-center retrospective case-control study including 253 consecutive adult patients undergoing surgery for confirmed or suspected primary fistula in ano who had undergone preoperative 3D-EAUS evaluation between January 2011 and January 2021. Main outcome measures were the concordance (K value) between ultrasound results and surgery in the identification of fistulas internal openings, primary tracts and secondary extensions and the 6 and 12 months success rate in patients with concordant and discordant findings. RESULTS: A good agreement in the identification of the main fistulas characteristics between ultrasound results and operative findings was found. A significant difference (P < .0001; Fisher's exact test) in the success rate was found between patients with concordant and discordant ultrasound results and operative findings in identification or location of internal opening. Particularly, all the 11 (4.8%) patients with discordant results experimented a failure of the surgical procedure at 6 months follow-up. At re-operation, the shift from discordant to concordant results was associated with an 81.8% 12 months success-rate. CONCLUSION: The three-dimensional endoanal ultrasound preoperative evaluation may have a relevant impact on the outcome of a defined group of patients undergoing surgery for anal fistula, since the careful evaluation of ultrasound results could simplify the internal orifice intra-operative detection and improve the success rate.

Cancer multidisciplinary team meetings: impact of logistical challenges on communication and decision-making.

BACKGROUND: Multidisciplinary teams (MDTs) are widely used in cancer care. Recent research points to logistical challenges impeding MDT decision-making and dissatisfaction among members. This study sought to identify different types of logistical issues and how they impacted team processes. METHODS: This was a secondary analysis of a cross-sectional observational study. Three cancer MDTs (breast, colorectal, and gynaecological) were recruited from UK hospitals. Validated observational instruments were used to measure decision-making (Metrics of Observational Decision-making, MDT-MODe), communication (Bales' Interaction Process Analysis, Bales' IPA), and case complexity (Measure of Case Discussion Complexity, MeDiC), including logistical challenges (Measure of Case Discussion Complexity, MeDiC), across 822 case discussions from 30 videoed meetings. Descriptive analysis and paired samples t tests were used to identify and compare frequency of different types of logistical challenges, along with partial correlations, controlling for clinical complexity of cases, to understand how such issues related to the MDT decision-making and communication. RESULTS: A significantly higher frequency of administrative and process issues (affecting 30 per cent of cases) was seen compared with the frequency of equipment issues (affecting 5 per cent of cases; P < 0.001) and the frequency of the attendance issues (affecting 16 per cent of cases; P < 0.001). The frequency of the attendance issues was significantly higher than the frequency of equipment issues (P < 0.001). Partial correlation analysis revealed that administrative and process issues, including attendance, were negatively correlated with quality of information (r = -0.15, P < 0.001; r = -0.11, P < 0.001), and equipment issues with the quality of contribution to meeting discussion (r = -0.14, P < 0.001). More questioning and answering by MDT members was evident with the administrative and process issues (r = 0.21, P < 0.001; r = 0.19, P < 0.001). Some differences were observed in teams' socioemotional reactions to the administrative and process issues with the gynaecological MDT showing positive correlation with positive socioemotional reactions (r = 0.20, P < 0.001), and the breast cancer MDT with negative socioemotional reactions (r = 0.17, P < 0.001). CONCLUSION: Administrative and process issues were the most frequent logistical challenges for the studied teams. Where diagnostic results were unavailable, and inadequate patient details provided, the quality of decision-making was reduced.

Transanal Hemorrhoidal Dearterialization (THD) Anolift-Prospective Assessment of Safety and Efficacy.

The adjunct of a mucopexy to conventional dearterialization has become a routine part of the transanal hemorrhoidal dearterialization procedure in order to facilitate the management of the prolapsing component and has helped to expand the indications of this technique to more advanced stages of hemorrhoidal disease. A simple technical modification of THD with targeted mucopexy (TM), called Anolift, is described. The aim of the study was to evaluate the safety and effectiveness of this technical variation. The procedure consisted of two parts: one aimed at the dearterialization and the other concentrated on the management of the prolapsing component. Once all the arteries were identified and transfixed an Anolift targeted mucopexy was performed using a continuous barbed suture with a synthetic absorbable monofilament (Polydioxanone) 2/0 Filbloc (Assut Europe) stitch mounted on a 4/8 30 mm needle. Severity of hemorrhoidal symptoms was scored from 0 to 20 using a dedicated questionnaire: the Hemorrhoidal Assessment Severity Score (HASS). From May 2018 to November 2020, 60 patients with hemorrhoidal disease (HD) underwent a THD Anolift procedure. Three patients experienced severe post-operative pain and 10 (23%) suffered with difficulty in evacuation. The median follow-up period was 15.5 months (range 2-32 months). The mean HASS changed from 16.43 pre-operatively to 1.95 post-operatively (p < 0.0001). Pre-operative HASS very strongly correlated with the degree of hemorrhoids (p < 0.001), while there was no correlation between the pre-operative HASS or the degree of hemorrhoids and the post-operative HASS (p = 0.163). There was no significant difference in predicted post-operative HASS according to the pre-operative HD stage. One patient (1.6%) with circumferential IV hemorrhoids had a recurrence and required a further THD. Two patients had excision of skin tags (3%). The Anolift technique is safe and effective for the management of HD even in patients with advanced stages.

A Biologic Surgical Implant in Complex Abdominal Wall Repair: 3-Year Follow-Up Results of a Multicentric Prospective Study.

PURPOSE: Despite the advancements in the reinforcement and closure techniques available, complex abdominal wall reconstruction (CAWR) remains a challenging surgical undertaking with considerable risk of postoperative complications. Biological meshes were developed that may help to complement standard closure techniques and offer an alternative to synthetic meshes, which carry significant risks with their use in complex cases. PATIENTS AND METHODS: A total of 114 patients underwent surgical treatment for CAWR with a Permacol™ (a biologic surgical implant). The study objective was to evaluate the short-term (6 months), mid-term (12-24 months), and long-term (36 months) clinical outcomes (through 36 months) associated with the use of a biologic surgical implant in these cases. RESULTS: The cumulative hernia recurrence rate was 18.7% (17/91) at 24 months and 22.4% (19/85) at 36 months. Twelve (14.1%) subjects required reoperation for hernia repair within 36 months for repair of recurrent hernias. Between 6- and 36-months post-surgery, patients reported improvement in their Carolina comfort scale (CSS) measures of severity of pain, sensation of mesh, and movement limitations. CONCLUSION: A biologic surgical implant can provide long-term benefit to complex abdominal wall repairs in patients staged grade III according to the Ventral Hernia Working Group (VHWG).

Chronic constipation in adults: Contemporary perspectives and clinical challenges. 2: Conservative, behavioural, medical and surgical treatment.

BACKGROUND: Chronic constipation is a prevalent disorder that affects quality of life of patients and consumes resources in healthcare systems worldwide. In clinical practice, it is still considered a challenge as clinicians frequently are unsure as to which treatments to use and when. Over a decade ago, a Neurogastroenterology and Motility journal supplement devoted to the investigation and management of constipation was published (Neurogastroenterol Motil 2009;21(Suppl 2):1). In October 2018, the 3rd London Masterclass, entitled "Contemporary management of constipation" was held. The faculty members of this symposium were invited to write two reviews to present a collective synthesis of talks presented and discussions held during this meeting. The first review addresses epidemiology, diagnosis, clinical associations, pathophysiology, and investigation. PURPOSE: The present is the second of these reviews, providing contemporary perspectives and clinical challenges regarding behavioral, conservative, medical, and surgical treatments for patients presenting with constipation. It includes a management algorithm to guide clinical practice.

A multicentre cross-sectional observational study of cancer multidisciplinary teams: Analysis of team decision making.

BACKGROUND: Multidisciplinary teams (MDT) formulate expert informed treatment recommendations for people with cancer. We set out to examine how the factors proposed by the functional perspective of group decision making (DM), that is, interaction process, internal factors (factors emanating from within the group such as group size), external circumstances (factors coming from the outside of the team), and case-complexity affect the quality of MDT decision making. METHODS: This was a cross-sectional observational study. Three cancer MDTs were recruited with 44 members overall and 30 of their weekly meetings filmed. Validated observational instruments were used to measure quality of DM, interactions, and complexity of 822 case discussions. RESULTS: The full regression model with the variables proposed by the functional perspective was significant, R2  = 0.52, F(20, 801) = 43.47, P < .001, adjusted R2  = 0.51. Positive predictors of DM quality were asking questions (P = .001), providing answers (P = .001), team size (P = .007), gender balance (P = .003), and clinical complexity (P = .001), while negative socioemotional reactions (P = .007), gender imbalance (P = .003), logistical issues (P = .001), time-workload pressures (P = .002), and time spent in the meeting (P = .001) were negative predictors. Second half of the meetings also saw significant decrease in the DM quality (P = .001), interactions (P = .001), group size (P = .003), and clinical complexity (P = .001), and an increase in negative socioemotional reactions (P = .001) and time-workload pressures (P = .001). DISCUSSION: To the best of our knowledge, this is the first study to attempt to assess the factors proposed by the functional perspective in cancer MDTs. One novel finding is the effect of sociocognitive factors on team DM quality, while another is the cognitive-catch 22 effect: while the case discussions are significantly simpler in the second half of the meeting, there is significantly less time left to discuss the remaining cases, further adding to the cognitive taxation in teams who are now rapidly attempting to close their time-workload gap. Implications are discussed in relation to quality and safety.

COVID-19 and the Global Impact on Colorectal Practice and Surgery.

BACKGROUND: The novel severe acute respiratory syndrome coronavirus 2 virus that emerged in December 2019 causing coronavirus disease 2019 (COVID-19) has led to the sudden national reorganization of health care systems and changes in the delivery of health care globally. The purpose of our study was to use a survey to assess the global effects of COVID-19 on colorectal practice and surgery. MATERIALS AND METHODS: A panel of International Society of University Colon and Rectal Surgeons (ISUCRS) selected 22 questions, which were included in the questionnaire. The questionnaire was distributed electronically to ISUCRS fellows and other surgeons included in the ISUCRS database and was advertised on social media sites. The questionnaire remained open from April 16 to 28, 2020. RESULTS: A total of 287 surgeons completed the survey. Of the 287 respondents, 90% were colorectal specialists or general surgeons with an interest in colorectal disease. COVID-19 had affected the practice of 96% of the surgeons, and 52% were now using telemedicine. Also, 66% reported that elective colorectal cancer surgery could proceed but with perioperative precautions. Of the 287 respondents, 19.5% reported that the use of personal protective equipment was the most important perioperative precaution. However, personal protective equipment was only provided by 9.1% of hospitals. In addition, 64% of surgeons were offering minimally invasive surgery. However, 44% reported that enough information was not available regarding the safety of the loss of intra-abdominal carbon dioxide gas during the COVID-19 pandemic. Finally, 61% of the surgeons were prepared to defer elective colorectal cancer surgery, with 29% willing to defer for ≤ 8 weeks. CONCLUSION: The results from our survey have demonstrated that, globally, COVID-19 has affected the ability of colorectal surgeons to offer care to their patients. We have also discussed suggestions for various practical adaptation strategies for use during the recovery period.

Biological Implant for Complex Abdominal Wall Reconstruction: A Single Institution Experience and Review of Literature.

AIM: To present our single institution and surgeon's complex abdominal wall reconstructions (CAWR) experience with Permacol™ mesh through a non-randomized study. PATIENTS AND METHOD: Data of 51 consecutive patients were prospectively collected between 2003 and 2015. Patients had a median of 3 comorbidities (range 0-10) and 68% were Center for Disease Control class II-IV. The mean previous repair was 1.3 (range, 0-12), and 25 (44%) had a mesh in situ. The median defect size was 625 cm2. RESULTS: Among the 56 CAWR procedures, in 16 (29%) bowel resection/anastomosis was performed. The overall post-operative complication rate was 45%, and it was wound-related except from 1 patient dead for myocardial infarction. One was lost at follow-up. Five were re-operated for recurrence with a second Permacol mesh, leading to 14 (26%) overall recurrences at a mean follow-up of 44 months (range, 4-123). In 33 (59%) cases, fascial closure was achieved. The mesh placement was intraperitoneal in 89%, retro-muscular in 9% and supra-fascial in 1% of cases. A multivariate analysis showed that predictor risk for recurrence was more than 3 previous repairs, wound class III-IV, whereas age, type of comorbidities, defect size and fascial closure did not influence the recurrence. Median post-operative performance status was 0 (range; 0-3). A satisfaction questionnaire was obtained in 43 patients, and 86% of them were satisfied with the outcome. CONCLUSIONS: Biological materials have the potential to reduce morbidity and improve outcome of definitive repair of CAWR.

The HubBLe Trial: haemorrhoidal artery ligation (HAL) versus rubber band ligation (RBL) for symptomatic second- and third-degree haemorrhoids: a multicentre randomised controlled trial and health-economic evaluation.

BACKGROUND: Optimal surgical intervention for low-grade haemorrhoids is unknown. Rubber band ligation (RBL) is probably the most common intervention. Haemorrhoidal artery ligation (HAL) is a novel alternative that may be more efficacious. OBJECTIVE: The comparison of HAL with RBL for the treatment of grade II/III haemorrhoids. DESIGN: A multicentre, parallel-group randomised controlled trial. PERSPECTIVE: UK NHS and Personal Social Services. SETTING: 17 NHS Trusts. PARTICIPANTS: Patients aged ≥ 18 years presenting with grade II/III (second- and third-degree) haemorrhoids, including those who have undergone previous RBL. INTERVENTIONS: HAL with Doppler probe compared with RBL. OUTCOMES: Primary outcome - recurrence at 1 year post procedure; secondary outcomes - recurrence at 6 weeks; haemorrhoid severity score; European Quality of Life-5 Dimensions, 5-level version (EQ-5D-5L); Vaizey incontinence score; pain assessment; complications; and cost-effectiveness. RESULTS: A total of 370 participants entered the trial. At 1 year post procedure, 30% of the HAL group had evidence of recurrence compared with 49% after RBL [adjusted odds ratio (OR) = 2.23, 95% confidence interval (CI) 1.42 to 3.51; p = 0.0005]. The main reason for the difference was the number of extra procedures required to achieve improvement/cure. If a single HAL is compared with multiple RBLs then only 37.5% recurred in the RBL arm (adjusted OR 1.35, 95% CI 0.85 to 2.15; p = 0.20). Persistence of significant symptoms at 6 weeks was lower in both arms than at 1 year (9% HAL and 29% RBL), suggesting significant deterioration in both groups over the year. Symptom score, EQ-5D-5L and Vaizey score improved in both groups compared with baseline, but there was no difference between interventions. Pain was less severe and of shorter duration in the RBL group; most of the HAL group who had pain had mild to moderate pain, resolving by 3 weeks. Complications were low frequency and not significantly different between groups. It appeared that HAL was not cost-effective compared with RBL. In the base-case analysis, the difference in mean total costs was £1027 higher for HAL. Quality-adjusted life-years (QALYs) were higher for HAL; however, the difference was very small (0.01) resulting in an incremental cost-effectiveness ratio of £104,427 per additional QALY. CONCLUSIONS: At 1 year, although HAL resulted in fewer recurrences, recurrence was similar to repeat RBL. Symptom scores, complications, EQ-5D-5L and continence score were no different, and patients had more pain in the early postoperative period after HAL. HAL is more expensive and unlikely to be cost-effective in terms of incremental cost per QALY. LIMITATIONS: Blinding of participants and site staff was not possible. FUTURE WORK: The incidence of recurrence may continue to increase with time. Further follow-up would add to the evidence regarding long-term clinical effectiveness and cost-effectiveness. The polysymptomatic nature of haemorrhoidal disease requires a validated scoring system, and the data from this trial will allow further assessment of validity of such a system. These data add to the literature regarding treatment of grade II/III haemorrhoids. The results dovetail with results from the eTHoS study [Watson AJM, Hudson J, Wood J, Kilonzo M, Brown SR, McDonald A, et al. Comparison of stapled haemorrhoidopexy with traditional excisional surgery for haemorrhoidal disease (eTHoS): a pragmatic, multicentre, randomised controlled trial. Lancet 2016, in press.] comparing stapled haemorrhoidectomy with excisional haemorrhoidectomy. Combined results will allow expansion of analysis, allowing surgeons to tailor their treatment options to individual patients. TRIAL REGISTRATION: Current Controlled Trials ISRCTN41394716. FUNDING: This project was funded by the NIHR Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 20, No. 88. See the NIHR Journals Library website for further project information.

Methionine adenosyltransferase α2 sumoylation positively regulate Bcl-2 expression in human colon and liver cancer cells.

Ubiquitin-conjugating enzyme 9 (Ubc9) is required for sumoylation and inhibits apoptosis via Bcl-2 by unknown mechanism. Methionine adenosyltransferase 2A (MAT2A) encodes for MATα2, the catalytic subunit of the MATII isoenzyme that synthesizes S-adenosylmethionine (SAMe). Ubc9, Bcl-2 and MAT2A expression are up-regulated in several malignancies. Exogenous SAMe decreases Ubc9 and MAT2A expression and is pro-apoptotic in liver and colon cancer cells. Here we investigated whether there is interplay between Ubc9, MAT2A and Bcl-2. We used human colon and liver cancer cell lines RKO and HepG2, respectively, and confirmed key finding in colon cancer specimens. We found MATα2 can regulate Bcl-2 expression at multiple levels. MATα2 binds to Bcl-2 promoter to activate its transcription. This effect is independent of SAMe as MATα2 catalytic mutant was also effective. MATα2 also directly interacts with Bcl-2 to enhance its protein stability. MATα2's effect on Bcl-2 requires Ubc9 as MATα2's stability is influenced by sumoylation at K340, K372 and K394. Overexpressing wild type (but not less stable MATα2 sumoylation mutants) protected from 5-fluorouracil-induced apoptosis in both colon and liver cancer cells. Colon cancer have higher levels of sumoylated MATα2, total MATα2, Ubc9 and Bcl-2 and higher MATα2 binding to the Bcl-2 P2 promoter. Taken together, Ubc9's protective effect on apoptosis may be mediated at least in part by sumoylating and stabilizing MATα2 protein, which in turn positively maintains Bcl-2 expression. These interactions feed forward to further enhance growth and survival of the cancer cell.

Successful combined management for ruptured superior rectal artery aneurysm in neurofibromatosis type 1.

Neurofibromatosis type 1 (NF1) is an autosomally dominant inherited disorder characterized by neurocutaneous and bony lesions with multisystem vasculopathy in the form of stenotic and aneurysmal disease. The multisystemic nature of NF1 aneurysmal disease can have potentially serious clinical sequelae and complicated management. A 55-year-old woman with NF1 presented acutely with severe abdominal and back pain, diarrhea, and hypotension with a delayed fall in her hemoglobin. Imaging demonstrated a retroperitoneal bleed from a superior rectal artery aneurysm. The inferior mesenteric artery was coiled with cessation of bleeding. She became unstable again and underwent a successful laparotomy for aneurysm exclusion and evacuation of hematoma. Vessel position and relatively nonspecific symptoms on presentation make awareness among front-line clinicians critical. Endovascular management can be difficult, and the clinician should be prepared for open surgery if the patient displays signs of recurrent bleeding.

Modern management of anal fistula.

Ideal surgical treatment for anal fistula should aim to eradicate sepsis and promote healing of the tract, whilst preserving the sphincters and the mechanism of continence. For the simple and most distal fistulae, conventional surgical options such as laying open of the fistula tract seem to be relatively safe and therefore, well accepted in clinical practise. However, for the more complex fistulae where a significant proportion of the anal sphincter is involved, great concern remains about damaging the sphincter and subsequent poor functional outcome, which is quite inevitable following conventional surgical treatment. For this reason, over the last two decades, many sphincter-preserving procedures for the treatment of anal fistula have been introduced with the common goal of minimising the injury to the anal sphincters and preserving optimal function. Among them, the ligation of intersphincteric fistula tract procedure appears to be safe and effective and may be routinely considered for complex anal fistula. Another technique, the anal fistula plug, derived from porcine small intestinal submucosa, is safe but modestly effective in long-term follow-up, with success rates varying from 24%-88%. The failure rate may be due to its extrusion from the fistula tract. To obviate that, a new designed plug (GORE BioA®) was introduced, but long term data regarding its efficacy are scant. Fibrin glue showed poor and variable healing rate (14%-74%). FiLaC and video-assisted anal fistula treatment procedures, respectively using laser and electrode energy, are expensive and yet to be thoroughly assessed in clinical practise. Recently, a therapy using autologous adipose-derived stem cells has been described. Their properties of regenerating tissues and suppressing inflammatory response must be better investigated on anal fistulae, and studies remain in progress. The aim of this present article is to review the pertinent literature, describing the advantages and limitations of new sphincter-preserving techniques.

A retrospective study evaluating the use of Permacol™ surgical implant in incisional and ventral hernia repair.

BACKGROUND: The outcome of incisional and ventral hernia repair depends on surgical technique, patient, and material. Permacol™ surgical implant (crosslinked porcine collagen) has been used for over a decade; however, there are few data on outcomes. This study is the largest retrospective multinational study to date to evaluate outcomes with Permacol™ surgical implant in the repair of incisional and ventral hernias. METHODS: Data were collected retrospectively on 343 patients treated for 213 incisional and 130 ventral hernias. Data evaluated included patient demographics, wound classification, surgical technique, morbidity, and recurrence rates. RESULTS: Median follow-up time was 649 days (max: 2857), median age 57 years (range 23-91), and BMI 32 kg/m(2) (range 17.6-77.8). Two or more comorbidities were present in 70% of patients. Open surgery was performed in 220 (64%) patients. Permacol™ surgical implant was used as an underlay (250), sublay (39), onlay (37), or inlay (17). Surgical techniques included component separation (89; 25.9%), modified Stoppa technique (197; 57.4%), and Rives-Stoppa (17; 5.0%). CDC Surgical Wound Classification was Class I (190), Class II (103), Class III (28), and Class IV (22). Complications were seen in 40.5% (139) of the patients, with seroma (19%) and wound infection (15%) as the most common. Mesh removal occurred in 1 (0.3%) patient. Kaplan-Meier analysis demonstrated that the probabilities for hernia recurrence at one, two, and three years were 5.8%, 16.6%, and 31.0%, respectively. CONCLUSIONS: Permacol™ surgical implant was shown to be safe with relatively low rates of hernia recurrence. CLINICAL TRIAL REGISTRATION NUMBER: NCT01214252 (http://www.clinicaltrials.gov).

Polyamine and methionine adenosyltransferase 2A crosstalk in human colon and liver cancer.

Methionine adenosyltransferase (MAT) is an essential enzyme that is responsible for the biosynthesis of S-adenosylmethionine (SAMe), the principal methyl donor and precursor of polyamines. MAT1A is expressed in normal liver and MAT2A is expressed in all extrahepatic tissues. MAT2A expression is increased in human colon cancer and in colon cancer cells treated with mitogens, whereas silencing MAT2A resulted in apoptosis. The aim of the current work was to examine the mechanism responsible for MAT2A-dependent growth and apoptosis. We found that in RKO (human adenocarcinoma cell line) cells, MAT2A siRNA treatment lowered cellular SAMe and putrescine levels by 70-75%, increased apoptosis and inhibited growth. Putrescine supplementation blunted significantly MAT2A siRNA-induced apoptosis and growth suppression. Putrescine treatment (100pmol/L) raised MAT2A mRNA level to 4.3-fold of control, increased the expression of c-Jun and c-Fos and binding to an AP-1 site in the human MAT2A promoter and the promoter activity. In human colon cancer specimens, the expression levels of MAT2A, ornithine decarboxylase (ODC), c-Jun and c-Fos are all elevated as compared to adjacent non-tumorous tissues. Overexpression of ODC in RKO cells also raised MAT2A mRNA level and MAT2A promoter activity. ODC and MAT2A are also overexpressed in liver cancer and consistently, similar MAT2A-ODC-putrescine interactions and effects on growth and apoptosis were observed in HepG2 cells. In conclusion, there is a crosstalk between polyamines and MAT2A. Increased MAT2A expression provides more SAMe for polyamines biosynthesis; increased polyamine (putrescine in this case) can activate MAT2A at the transcriptional level. This along with increased ODC expression in cancer all feed forward to further enhance the proliferative capacity of the cancer cell.

MAT2B-GIT1 interplay activates MEK1/ERK 1 and 2 to induce growth in human liver and colon cancer.

UNLABELLED: Methionine adenosyltransferase 2B (MAT2B) encodes for two variant proteins (V1 and V2) that promote cell growth. Using in-solution proteomics, GIT1 (G Protein Coupled Receptor Kinase Interacting ArfGAP 1) was identified as a potential interacting partner of MAT2B. Here, we examined the functional significance of this interplay. Coimmunoprecipitation experiments examined protein interactions. Tissue expression levels of proteins were examined using immunohistochemistry and western blotting. Expression levels of proteins were varied using transient knockdown or overexpression to observe the effect of alterations in each protein on the entire complex. Direct interaction among individual proteins was further verified using in vitro translated and recombinant proteins. We found both MAT2B variants interact with GIT1. Overexpression of V1, V2, or GIT1 activated mitogen-activated protein kinase kinase 1 (MEK1) and extracellular signal-regulated kinase (ERK), raised cyclin D1 protein level, and increased growth, whereas the opposite occurred when V1, V2, or GIT1 was knocked down. MAT2B and GIT1 require each other to activate MEK1/ERK and increase growth. MAT2B directly interacts with MEK1, GIT1, and ERK2. Expression level of V1, V2, or GIT1 directly influenced recruitment of GIT1 or MAT2B and ERK2 to MEK1, respectively. In pull-down assays, MAT2B directly promoted binding of GIT1 and ERK2 to MEK1. MAT2B and GIT1 interact and are overexpressed in most human liver and colon cancer specimens. Increased expression of V1, V2, or GIT1 promoted growth in an orthotopic liver cancer model, whereas increased expression of either V1 or V2 with GIT1 further enhanced growth and lung metastasis. CONCLUSION: MAT2B and GIT1 form a scaffold, which recruits and activates MEK and ERK to promote growth and tumorigenesis. This novel MAT2B/GIT1 complex may provide a potential therapeutic gateway in human liver and colon cancer. (HEPATOLOGY 2012).

S-adenosyl methionine regulates ubiquitin-conjugating enzyme 9 protein expression and sumoylation in murine liver and human cancers.

UNLABELLED: Ubiquitin-conjugating enzyme 9 (Ubc9) is required for sumoylation and is overexpressed in several malignancies, but its expression in hepatocellular carcinoma (HCC) is unknown. Hepatic S-adenosyl methionine (SAMe) levels decrease in methionine adenosyltransferase 1A (Mat1a) knockout (KO) mice, which develop HCC, and in ethanol-fed mice. We examined the regulation of Ubc9 by SAMe in murine liver and human HCC, breast, and colon carcinoma cell lines and specimens. Real-time polymerase chain reaction and western blotting measured gene and protein expression, respectively. Immunoprecipitation followed by western blotting examined protein-protein interactions. Ubc9 expression increased in HCC and when hepatic SAMe levels decreased. SAMe treatment in Mat1a KO mice reduced Ubc9 protein, but not messenger RNA (mRNA) levels, and lowered sumoylation. Similarly, treatment of liver cancer cell lines HepG2 and Huh7, colon cancer cell line RKO, and breast cancer cell line MCF-7 with SAMe or its metabolite 5'-methylthioadenosine (MTA) reduced only Ubc9 protein level. Ubc9 posttranslational regulation is unknown. Ubc9 sequence predicted a possible phosphorylation site by cell division cycle 2 (Cdc2), which directly phosphorylated recombinant Ubc9. Mat1a KO mice had higher phosphorylated (phospho)-Ubc9 levels, which normalized after SAMe treatment. SAMe and MTA treatment lowered Cdc2 mRNA and protein levels, as well as phospho-Ubc9 and protein sumoylation in liver, colon, and breast cancer cells. Serine 71 of Ubc9 was required for phosphorylation, interaction with Cdc2, and protein stability. Cdc2, Ubc9, and phospho-Ubc9 levels increased in human liver, breast, and colon cancers. CONCLUSION: Cdc2 expression is increased and Ubc9 is hyperphosphorylated in several cancers, and this represents a novel mechanism to maintain high Ubc9 protein expression that can be inhibited by SAMe and MTA.