MCCC2 promotes HCC development by supporting leucine oncogenic function.

BACKGROUND: The role of methylcrotonoyl-CoA carboxylase 2 (MCCC2) in the development of tumors is well-established, and the involvement of leucine in the liver is well-known. However, the role of MCCC2 and the correlation between MCCC2 and leucine in the progression of hepatocellular carcinoma (HCC) have not yet been reported. METHODS: In this study, the Gepia database was used to evaluate the prognostic value of MCCC2 in HCC. The expression and localization of MCCC2 in HCC cells were determined by western blot and immunofluorescence assays. Flow cytometry and CCK-8 and transwell assays were carried out to explore the effect of MCCC2 on cell proliferation, migration, and invasion. In addition, mass spectrometry analysis was used to predict the potential cell function of MCCC2 in HCC. RESULTS: We found that the expression of MCCC2 increased in HCC tissues and that high expression of MCCC2 could predict poor outcomes in HCC patients. Knockdown expression of MCCC2 in HCC cells could reduce cell proliferation, migration, and invasion ability in vitro and could inhibit HCC cell proliferation in vivo. Interestingly, we found that HCC cells transfected with MCCC2-sgRNA failed to respond to leucine deprivation. Meanwhile, leucine deprivation inhibited cell proliferation, migration, and invasion in HCC cells where MCCC2 was present rather than in cells where MCCC2 was absent. In addition, knockdown of MCCC2 significantly reduced the glycolysis markers, glucose consumption, lactate secretion, and acetyl-CoA level, which is a product of leucine metabolism. Furthermore, we found that MCCC2 promotes the activation of ERK. Profiling the MCCC2 binding proteins revealed that MCCC2-associated proteins are enriched in biological processes, such as protein metabolism, energy pathway, and metabolism in HCC cells. CONCLUSIONS: Our findings revealed that MCCC2 plays a critical role in the development of HCC, and the leucine metabolism pathway might be a novel target in HCC treatment.

The AKR1B1 inhibitor epalrestat suppresses the progression of cervical cancer.

Cervical cancer is the leading cause of cancer-related death among women worldwide. Identifying an effective treatment with fewer side effects is imperative, because all of the current treatments have unique disadvantages. Aldo-keto reductase family 1 member B1 (AKR1B1) is highly expressed in various cancers and is associated with tumor development, but has not been studied in cervical cancer. In the current study, we used CRISPR/Cas9 technology to establish a stable HeLa cell line with AKR1B1 knockout. In vitro, AKR1B1 knockout inhibited the proliferation, migration and invasion of HeLa cells, providing evidence that AKR1B1 is an innovative therapeutic target. Notably, the clinically used epalrestat, an inhibitor of aldose reductases, including AKR1B1, had the same effect as AKR1B1 knockout on HeLa cells. This result suggests that epalrestat could be used in the clinical treatment of cervical cancer, a prospect that undoubtedly requires further research. Moreover, aiming to determine the underlying regulatory mechanism of AKR1B1, we screened a series of differentially regulated genes (DEGs) by RNA sequencing and verified selected DEGs by quantitative RT-PCR. In addition, gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of the DEGs revealed a correlation between AKR1B1 and cancer. In summary, epalrestat inhibits the progression of cervical cancer by inhibiting AKR1B1, and thus may be a new drug for the clinical treatment of cervical cancer.

Correlation between S100A11 and the TGF-ß1/SMAD4 pathway and its effects on the proliferation and apoptosis of pancreatic cancer cell line PANC-1.

S100A11 as a S100 protein family member has been documented to play dual-direction regulation over cancer cell proliferation. We explored the role of S100A11 in the proliferation and apoptosis of pancreatic cancer cell line PANC-1 and the potential mechanisms involving the TGF-ß1/SMAD4/p21 pathway. S100A11 and TGF-ß1 protein expressions in 30 paraffin-embedded specimens were evaluated by immunohistochemistry. S100A11 and TGF-ß1 expression in PANC-1 cell line was suppressed using small interfering RNA (siRNA), respectively. Subsequently, pancreatic cancer cell apoptosis was measured by Cell Counting Kit-8 and flow cytometry, and S100A11 and TGF-ß1/SMAD4/p21 pathway proteins and genes were detected with Western blotting and quantitative polymerase chain reaction (qPCR). S100A11 cytoplasmic/nuclear protein translocation was examined using NE-PER® cytoplasm/nuclear protein extraction in cells interfered with TGF-ß1 siRNA. Our results showed that S100A11 expression was positively correlated with TGF-ß1 expression in pancreatic cancerous tissue. Silencing TGF-ß1 down-regulated intracellular P21WAF1 expression by 90%, blocked S100A11 from cytoplasm entering nucleus, and enhanced cell proliferation. Silencing S100A11 down-regulated intracellular P21 expression and promoted cell apoptosis without significantly changing TGF-ß1 and SMAD4 expression. Our findings revealed that S100A11 and TGF-ß1/SMAD4 signaling pathway were related but mutually independent in regulating PANC-1 cells proliferation and apoptosis. Other independent mechanisms might be involved in S100A11's regulation of pancreatic cell growth. S100A11 could be a potential gene therapy target for pancreatic cancer.

Interleukin-34 deficiency aggravates development of colitis and colitis-associated cancer in mice.

BACKGROUND: Although expression of interleukin (IL)-34 is upregulated in active ulcerative colitis (UC), the molecular function and underlying mechanism are largely unclear. AIM: To investigate the function of IL-34 in acute colitis, in a wound healing model and in colitis-associated cancer in IL-34-deficient mice. METHODS: Colitis was induced by administration of dextran sodium sulfate (DSS), and carcinogenesis was induced by azoxymethane (AOM). Whether the impact of IL-34 on colitis was dependent on macrophages was validated by depletion of macrophages in a murine model. The association between IL-34 expression and epithelial proliferation was studied in patients with active UC. RESULTS: IL-34 deficiency aggravated murine colitis in acute colitis and in wound healing phase. The effect of IL-34 on experimental colitis was not dependent on macrophage differentiation and polarization. IL-34-deficient mice developed more tumors than wild-type mice following administration of AOM and DSS. No significant difference was shown in degree of cellular differentiation in tumors between wild-type and IL-34-deficient mice. IL-34 was dramatically increased in the active UC patients as previously reported. More importantly, expression of IL-34 was positively correlated with epithelial cell proliferation in patients with UC. CONCLUSION: IL-34 deficiency exacerbates colonic inflammation and accelerates colitis-associated carcinogenesis in mice. It might be served as a potential therapeutic target in UC.

Diagnosis and management of dysplasia and cancer of the ileal pouch in patients with underlying inflammatory bowel disease.

Approximately 30% of the patients with ulcerative colitis (UC) would ultimately require colectomy for medically refractory UC or UC-associated neoplasia. Restorative proctocolectomy with ileal pouch-anal anastomosis has become the surgical treatment of choice for these patients. However, this procedure does not completely abolish the risk for neoplasia of the pouch. The main risk factor for pouch neoplasia is a preoperative diagnosis of UC-associated dysplasia or cancer. Although the natural history and prognosis of pouch dysplasia are not clear, mortality associated with pouch cancer, once diagnosed, appears to be high. Conversely, not all pouch neoplasia follows the chronic inflammation-dysplasia-cancer sequence, which makes pouch endoscopy with biopsy, the current gold standard for surveillance, challenging. In addition, the findings that pouch neoplasia is not common and that pouch endoscopy still misses dysplasia lead to controversy on the need and time interval of routine endoscopic surveillance. However, based on reports in the literature and their own experience, the authors recommend surveillance endoscopy to be performed in patients at risk, such as those with a precolectomy diagnosis of UC-associated neoplasia. This review appraises issues in the prevalence and incidence, risk factors, technical aspects of pouch construction, clinical and pathological features, natural history, surveillance examination, diagnosis, and management of pouch neoplasia.

Transmural inflammation is not pathognomonic for Crohn's disease of the pouch.

BACKGROUND: Transmural inflammation shown by imaging and histology has been considered a hallmark of Crohn's disease (CD). However, the diagnostic and prognostic value of this feature in CD of the pouch has not been evaluated. This study aimed to evaluate the clinical utility of transmural inflammation in patients with ileal pouch-anal anastomosis (IPAA) using in vivo optical coherence tomography (OCT) and histopathology. METHODS: All the patients were recruited from the subspecialty Pouchitis Clinic. The study consisted of two parts: (1) a prospective study with in vivo through-the-scope OCT for the evaluation of transmural disease in patients with normal or diseased pouches and (2) a retrospective pathology re-review for transmural inflammation in excised pouch specimens of CD and chronic pouchitis. RESULTS: This prospective OCT study enrolled 53 patients: 11 (20.8%) with normal pouches or irritable pouch syndrome, 10 (18.9%) with acute pouchitis, 11 (20.8%) with chronic antibiotic-refractory pouchitis (CARP), and 21 (39.6%) with CD of the pouch. Transmural inflammation, characterized by the loss of layered structure on OCT, was detected in 16 patients (30.2%): 4 with chronic pouchitis and 12 with CD of the pouch. None of the patients with normal pouches, irritable pouch syndrome, or acute pouchitis had transmural disease shown on OCT. Of the 26 patients with pouch failure who had pouch excision, the surgical specimens showed transmural disease in 30% of the CARP patients (3/10) and 12.5% (2/16) of those with CD of the pouch. CONCLUSIONS: Transmural disease in the setting of IPAA is not pathognomonic of CD. Transmural inflammation shown by imaging or histopathology was seen in both CD and CARP. Transmural inflammation of the pouch appeared to be associated with poor pouch outcome.

High expression of ubiquitin-conjugating enzyme E2A predicts poor prognosis in hepatocellular carcinoma.

The present study aimed to illustrate the association of the expression of ubiquitin-conjugating enzyme E2A (UBE2A) with the clinicopathological parameters and prognosis in hepatocellular carcinoma (HCC). The expression levels of UBE2A mRNA and protein in a total of 276 HCC tissues and six liver cell lines was detected by fluorescent quantitative polymerase chain reaction, western blotting and immunohistochemistry. Statistical analysis was also performed to assess the association of the expression of UBE2A with the clinicopathological parameters and prognosis by the GraphPad Prism and SPSS version 21.0 software. UBE2A mRNA and protein were highly expressed in HCC tissues compared with those in the adjacent normal tissue. Immunohistochemical analysis revealed that UBE2A protein was more strongly stained in the 276 paraffin-embedded HCC tissues as compared with the 63 adjacent normal tissue. Statistical analysis also demonstrated that UBE2A expression was significantly associated with histological differentiation, TNM stage and vascular invasion of HCC (P<0.05). Notably, HCC patients with a high expression of UBE2A had a shorter survival time as compared with those with a low expression of UBE2A. There results suggested that UBE2A may be involved in the pathogenesis of HCC and may serve as an important prognostic marker. Further exploration of the involvement of UBE2A in HCC development may provide novel therapeutic targets.

Transplantation of mouse embryonic stem cell-derived oligodendrocytes in the murine model of globoid cell leukodystrophy.

INTRODUCTION: Globoid cell leukodystrophy (GLD) is a severe disorder of the central and peripheral nervous system caused by the absence of galactocerebrosidase (GALC) activity. Cell-based therapies are highly promising strategies for GLD. In this study, G-Olig2 mouse embryonic stem cells (ESCs) were induced into oligodendrocyte progenitor cells (OPCs) and were implanted into the brains of twitcher mice, an animal model of GLD, to explore the therapeutic potential of the cells. METHODS: The G-Olig2 ESCs were induced into OPCs by using cytokines and a multi-step differentiation procedure. Oligodendrocyte markers were detected by reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry. The toxicity of psychosine to OPCs was determined by a cell proliferation assay kit. The GALC level of OPCs was also examined. OPCs were labeled with Dir and transplanted into the brains of twitcher mice. The transplanted cells were detected by in-Vivo Multispectral Imaging System and real-time PCR. The physiological effects of twitcher mice were assessed. RESULTS: Oligodendrocyte markers were expressed in OPCs, and 76%±5.76% of the OPCs were enhanced green fluorescent protein (eGFP)-positive, eGFP was driven by the Olig2 promoter. The effect of psychosine on cell viability indicated that OPCs were more resistant to psychosine toxicity. The GALC level of OPCs was 10.0±1.23 nmol/hour per mg protein, which was significantly higher than other cells. Dir-labeled OPCs were injected into the forebrain of post-natal day 10 twitcher mice. The transplanted OPCs were myelin basic protein (MBP)-positive and remained along the injection tract as observed by fluorescent microscopy. The level of the Dir fluorescent signal and eGFP mRNA significantly decreased at days 10 and 20 after injection, as indicated by in-Vivo Multispectral Imaging System and real-time PCR. Because of poor cell survival and limited migration ability, there was no significant improvement in brain GALC activity, MBP level, life span, body weight, and behavioral deficits of twitcher mice. CONCLUSIONS: ESC-derived OPC transplantation was not sufficient to reverse the clinical course of GLD in twitcher mice.

Chronic pouchitis is associated with pouch polyp formation in patients with underlying ulcerative colitis.

BACKGROUND: Polypoid lesions can develop in ileal pouches. The risk factors associated with the development of pouch polyps have not been studied. AIM: To characterize clinical features, risk factors, and disease course of pouch polyp in a cohort of patients with underlying inflammatory bowel disease (IBD) from a subspecialty clinic. METHOD: A total of 1094 patients with restorative proctocolectomy and IPAA for IBD presenting to our Pouchitis Clinic from 2002 to 2010 were included. Demographic, clinical, and endoscopic variables were analyzed. RESULTS: The median durations from UC diagnosis to colectomy and from pouch creation to the last follow-up for the whole cohort were 6 (interquartile range [IQR]: 3-13) and 9years (IQR: 5-14), respectively. A total of 2472 surveillance and/or diagnostic pouchoscopies were performed for the cohort with a median follow-up of 5 (IQR: 2-6) years in the Pouchitis Clinic. The median number of pouchoscopies per patient was 2 (IQR: 1-3). Of the 1094 patients, 96 (8.8%) were found to have pouch polyps. The median size of the polyps was 1.2 (IQR: 1.0-2.0) cm. On histology, 93 patients (96.9%) had inflammatory-type polyps and 3 (3.1%) had polyps with low-grade dysplasia or indefinite for dysplasia. Multivariate logistic regression analysis demonstrated that chronic pouch inflammatory change was a risk factor for the development of pouch polyp with an odds ratio of 2.26 (95% confidence interval: 1.35-3.79; P=0.002). CONCLUSION: The majority of pouch polyps in patients with underlying UC were benign. Patients with concomitant chronic pouch inflammatory changes had an increased risk for developing pouch polyps.

Clinical significance of indefinite for dysplasia on pouch biopsy in patients with underlying inflammatory bowel disease.

BACKGROUND: "Indefinite for dysplasia" (IND) on pouch mucosal biopsy is occasionally reported during routine histopathological evaluation. The natural history and implication of this histologic entity in ileal pouch-anal anastomosis (IPAA) has not been studied. AIM: The aim of this study is to characterize cumulative probability, natural history, and clinical outcome of pouch IND in a cohort of patients with inflammatory bowel disease (IBD). METHODS: All 932 patients with restorative proctocolectomy and IPAA for IBD were included. Patients with or without IND were classified into the study and control groups. Demographic, clinical, endoscopic, and histologic variables were analyzed. RESULTS: The mean duration from IBD diagnosis to colectomy and from pouch construction to data entry was 8.4 ± 8.5 and 9.7 ± 6.2 years, respectively. A total of 2,250 surveillance or diagnostic pouchoscopies with biopsies were performed for the cohort. Twenty-one patients (2.3%) were diagnosed with anal transitional zone and/or pouch IND, for whom subsequent pouchoscopies were performed with the mean procedure number being 3.4 ± 2.2 per patient during a mean of follow-up of 19.3 ± 16.1 months. One patient with IND developed low-grade dysplasia and one had high-grade dysplasia in a separate endoscopy. Cox model showed the presence of primary sclerosing cholangitis was an independent risk factor for pouch IND [hazard ratio = 6.76 (95% CI 2.56-17.88)]. Interobserver agreement (kappa score) for diagnosing pouch IND between GI pathologists ranged from 0.67 to 0.76. CONCLUSIONS: Subsequent dysplasia was uncommon in pouch patients with IND. Natural history of pouch IND warrants further long-term investigation.