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Background: Despite consensus supporting enhanced recovery programs, their full implementation in such a context is difficult due to conventional practices within various groups of professionals. The goal of the EUropean PErioperative MEdical Networking (EUPEMEN) project was to bring together the expertise and experience of national clinical professionals who have previously helped deliver major change programs in their countries and to use them to spread enhanced recovery after surgery protocols (ERAS) in Europe. The specific aim of this study is to present and discuss the key points of the proposed recommendations for colorectal surgery., Materials and Methods: Five partners from university hospitals in four European countries developed the project as partners. Following a non-systematic review of the literature, the European consensus panel generated a list of recommendations for perioperative care in colorectal surgery. A list of recommendations was formulated and distributed to collaborators at each center to allow modifications or additional statements. These recommendations were then discussed in three consecutive meetings to share uniform ERAS protocols to be disseminated., Result: The working group developed (1) the EUPEMEN online platform to offer, free of charge, evidence-based standardized perioperative care protocols, learning activities, and assistance to health professionals interested in enhancing the recovery of their patients; (2) the preparation of the EUPEMEN Multimodal Rehabilitation manuals; (3) the training of the trainers to teach future teachers; and (4) the dissemination of the results in five multiplier events, one for each partner, to promote and disseminate the protocols., Conclusion: The EUPEMEN project allowed the sharing of the expertise of many professionals from four different European countries with the objective of training the new generations in the dissemination of ERAS protocols in daily clinical practice through a new learning system. This project was proposed as an additional training tool for all the enhanced recovery program teams., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Unlabelled: Down-regulation of the liver-specific MAT1A gene, encoding S-adenosylmethionine (SAM) synthesizing isozymes MATI/III, and up-regulation of widely expressed MAT2A, encoding MATII isozyme, known as MAT1A:MAT2A switch, occurs in hepatocellular carcinoma (HCC). Here we found Mat1A:Mat2A switch and low SAM levels, associated with CpG hypermethylation and histone H4 deacetylation of Mat1A promoter, and prevalent CpG hypomethylation and histone H4 acetylation in Mat2A promoter of fast-growing HCC of F344 rats, genetically susceptible to hepatocarcinogenesis. In HCC of genetically resistant BN rats, very low changes in the Mat1A:Mat2A ratio, CpG methylation, and histone H4 acetylation occurred. The highest MAT1A promoter hypermethylation and MAT2A promoter hypomethylation occurred in human HCC with poorer prognosis. Furthermore, levels of AUF1 protein, which destabilizes MAT1A messenger RNA (mRNA), Mat1A-AUF1 ribonucleoprotein, HuR protein, which stabilizes MAT2A mRNA, and Mat2A-HuR ribonucleoprotein sharply increased in F344 and human HCC, and underwent low/no increase in BN HCC. In human HCC, Mat1A:MAT2A expression and MATI/III:MATII activity ratios correlated negatively with cell proliferation and genomic instability, and positively with apoptosis and DNA methylation. Noticeably, the MATI/III:MATII ratio strongly predicted patient survival length. Forced MAT1A overexpression in HepG2 and HuH7 cells led to a rise in the SAM level, decreased cell proliferation, increased apoptosis, down-regulation of Cyclin D1, E2F1, IKK, NF-κB, and antiapoptotic BCL2 and XIAP genes, and up-regulation of BAX and BAK proapoptotic genes. In conclusion, we found for the first time a post-transcriptional regulation of MAT1A and MAT2A by AUF1 and HuR in HCC. Low MATI/III:MATII ratio is a prognostic marker that contributes to determine a phenotype susceptible to HCC and patients' survival., Conclusion: Interference with cell cycle progression and I-kappa B kinase (IKK)/nuclear factor kappa B (NF-κB) signaling contributes to the antiproliferative and proapoptotic effect of high SAM levels in HCC., (Copyright © 2012 American Association for the Study of Liver Diseases.)

Unlabelled: The identification of molecular mechanisms involved in the maintenance of the transformed phenotype of hepatocellular carcinoma (HCC) cells is essential for the elucidation of therapeutic strategies. Here, we show that human HCC cells display an autocrine loop mediated by connective tissue growth factor (CTGF) that promotes DNA synthesis and cell survival. Expression of CTGF was stimulated by epidermal growth factor receptor (EGFR) ligands and was dependent on the expression of the transcriptional coactivator, Yes-associated protein (YAP). We identified elements in the CTGF gene proximal promoter that bound YAP-enclosing complexes and were responsible for basal and EGFR-stimulated CTGF expression. We also demonstrate that YAP expression can be up-regulated through EGFR activation not only in HCC cells, but also in primary human hepatocytes. CTGF contributed to HCC cell dedifferentiation, expression of inflammation-related genes involved in carcinogenesis, resistance toward doxorubicin, and in vivo HCC cell growth. Importantly, CTGF down-regulated tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor 2 expression and was involved in the reduced sensitivity of these cells toward TRAIL-mediated apoptosis., Conclusion: We have identified autocrine CTGF as a novel determinant of HCC cells' neoplastic behavior. Expression of CTGF can be stimulated through the EGFR-signaling system in HCC cells in a novel cross-talk with the oncoprotein YAP. Moreover, to our knowledge, this is the first study that identifies a signaling mechanism triggering YAP gene expression in healthy and transformed liver parenchymal cells., (Copyright © 2011 American Association for the Study of Liver Diseases.)

Unlabelled: Up-regulation of the v-Myb avian myeloblastosis viral oncogene homolog-like2 B-Myb (MYBL2) gene occurs in human hepatocellular carcinoma (HCC) and is associated with faster progression of rodent hepatocarcinogenesis. We evaluated, in distinct human HCC prognostic subtypes (as defined by patient survival length), activation of MYBL2 and MYBL2-related genes, and relationships of p53 status with MYBL2 activity. Highest total and phosphorylated protein levels of MYBL2, E2F1-DP1, inactivated retinoblastoma protein (pRB), and cyclin B1 occurred in HCC with poorer outcome (HCCP), compared to HCC with better outcome (HCCB). In HCCP, highest LIN9-MYBL2 complex (LINC) and lowest inactive LIN9-p130 complex levels occurred. MYBL2 positively correlated with HCC genomic instability, proliferation, and microvessel density, and negatively with apoptosis. Higher MYBL2/LINC activation in HCC with mutated p53 was in contrast with LINC inactivation in HCC harboring wildtype p53. Small interfering RNA (siRNA)-mediated MYBL2/LINC silencing reduced proliferation, induced apoptosis, and DNA damage at similar levels in HCC cell lines, irrespective of p53 status. However, association of MYBL2/LINC silencing with doxorubicin-induced DNA damage caused stronger growth restraint in p53(-/-) Huh7 and Hep3B cells than in p53(+/+) Huh6 and HepG2 cells. Doxorubicin triggered LIN9 dissociation from MYBL2 in p53(+/+) cell lines and increased MYBL2-LIN9 complexes in p53(-/-) cells. Doxorubicin-induced MYBL2 dissociation from LIN9 led to p21(WAF1) up-regulation in p53(+/+) but not in p53(-/-) cell lines. Suppression of p53 or p21(WAF1) genes abolished DNA damage response, enhanced apoptosis, and inhibited growth in doxorubicin-treated cells harboring p53(+/+) ., Conclusion: We show that MYBL2 activation is crucial for human HCC progression. In particular, our data indicate that MYBL2-LIN9 complex integrity contributes to survival of DNA damaged p53(-/-) cells. Thus, MYBL2 inhibition could represent a valuable adjuvant for treatments against human HCC with mutated p53., (2011 American Association for the Study of Liver Diseases.)

Unlabelled: Increased mitogen-activated protein kinase (MAPK) activity correlates with a more malignant hepatocellular carcinoma (HCC) phenotype. There is a reciprocal regulation between p44/42 MAPK (extracellular signal-regulated kinase [ERK]1/2) and the dual-specificity MAPK phosphatase MKP-1/DUSP1. ERK phosphorylates DUSP1, facilitating its proteasomal degradation, whereas DUSP1 inhibits ERK activity. Methionine adenosyltransferase 1a (Mat1a) knockout (KO) mice express hepatic S-adenosylmethionine (SAM) deficiency and increased ERK activity and develop HCC. The aim of this study was to examine whether DUSP1 expression is regulated by SAM and if so, elucidate the molecular mechanisms. Studies were conducted using Mat1a KO mice livers, cultured mouse and human hepatocytes, and 20S and 26S proteasomes. DUSP1 messenger RNA (mRNA) and protein levels were reduced markedly in livers of Mat1a KO mice and in cultured mouse and human hepatocytes with protein falling to lower levels than mRNA. SAM treatment protected against the fall in DUSP1 mRNA and protein levels in mouse and human hepatocytes. SAM increased DUSP1 transcription, p53 binding to DUSP1 promoter, and stability of its mRNA and protein. Proteasomal chymotrypsin-like and caspase-like activities were increased in Mat1a KO livers and cultured hepatocytes, which was blocked by SAM treatment. SAM inhibited chymotrypsin-like and caspase-like activities by 40% and 70%, respectively, in 20S proteasomes and caused rapid degradation of some of the 26S proteasomal subunits, which was blocked by the proteasome inhibitor MG132. SAM treatment in Mat1a KO mice for 7 days raised SAM, DUSP1, mRNA and protein levels and lowered proteosomal and ERK activities., Conclusion: DUSP1 mRNA and protein levels are lower in Mat1a KO livers and fall rapidly in cultured hepatocytes. SAM treatment increases DUSP1 expression through multiple mechanisms, and this may suppress ERK activity and malignant degeneration.

Unlabelled: The genome of the BALB/c mouse strain provides alleles that dominantly inhibit hepatocellular tumor development in F1 crosses with the highly hepatocarcinogenesis-susceptible C3H/He strain. Genome-wide linkage analysis using a 1536-single-nucleotide polymorphism array in a (C3H/He x BALB/c)F2 intercross population treated with urethane to induce hepatocellular tumor development revealed a locus with a major role in the resistance to hepatocarcinogenesis. This locus, designated hepatocarcinogen resistance 3 (Hpcr3) and mapping to central chromosome 15, showed a linkage at LOD score = 16.52 and accounted for 40% of the phenotypical variance. The BALB/c-derived allele at Hpcr3 reduced tumor-occupied area of the liver up to 25-fold, in a semidominant way. Additional minor loci were mapped to chromosomes 1, 10, and 18. A gene expression profile of normal adult mouse liver showed a significant association with susceptibility of BALB/c, C3H/He, and F1 mice to hepatocarcinogenesis and identified the genes expressed in the Hpcr3 locus region; moreover, this analysis implicated the E2F1 pathway in the modulation of the phenotype susceptibility to hepatocarcinogenesis., Conclusion: These findings, indicating the complex genetics of dominant resistance to hepatocarcinogenesis, represent a step toward the identification of the genes underlying this phenotype.

Purpose: To evaluate the correlation between polarimetry parameters and standard threshold perimetry indices., Patient and Methods: One hundred and eight consecutive glaucomatous patients were recruited in this study. Glaucomatous patients were classified when an abnormal visual field and/or an abnormal optic nerve head were present. One eye was chosen randomly from all the subjects. All optic nerve heads were examined with the GDx variable corneal compensator (VCC) (Laser Diagnostic Technologies, Inc, San Diego, CA) and visual fields were assessed by Humphrey Perimeter, program 24-2, SITA standard (Humphrey Instrument, Inc, San Leandro, CA). GDx VCC parameters and visual field indices were considered. Correlation coefficient was used to compare the 2 sets of data. A linear regression model was also used to calculate the independent contribution of each GDx VCC parameter and to determine which parameter was the most predictive of visual field damage., Results: The mean age (+/-standard deviation) was 61+/-10.3, the average of the mean deviation (MD) was -2.31+/-3.2 dB and the mean pattern standard deviation (PSD) was 3.32+/-2.83 dB. Significant (P<0.05) correlation was found between MD and the nerve fiber indicator (NFI) (r=-0.35), the superior average (r=0.28), the Temporal-Superior Nasal-Inferior Temporal average (r=0.24) and the inferior average (r=0.21). The linear regression model found NFI to be the predictor variable of MD. Significant (P<0.05) correlation was found between PSD and the NFI (r=0.39), the superior and the inferior average (r=0.30), and the Temporal-Superior Nasal-Inferior Temporal average (r=0.29). The linear regression model found NFI to be the predictor variable of PSD., Conclusions: Some of the GDx VCC indices were significantly correlated to visual field indices in glaucomatous patients, but NFI was the best indicator of visual field damage.

Current evidence indicates that neoplastic nodules induced in liver of Brown Norway (BN) rats genetically resistant to hepatocarcinogenesis are not prone to evolve into hepatocellular carcinoma. We show that BN rats subjected to diethylnitrosamine/2-acetylaminofluorene/partial hepatectomy treatment with a "resistant hepatocyte" protocol displayed higher number of glutathione-S-transferase 7-7(+) hepatocytes when compared with susceptible Fisher 344 (F344) rats, both during and at the end of 2-acetylaminofluorene treatment. However, DNA synthesis declined in BN but not F344 rats after completion of reparative growth. Upregulation of p16(INK4A), Hsp90, and Cdc37 genes; an increase in Cdc37-Cdk4 complexes; and a decrease in p16(INK4A)-Cdk4 complexes occurred in preneoplastic liver, nodules, and hepatocellular carcinoma of F344 rats. These parameters did not change significantly in BN rats. E2f4 was equally expressed in the lesions of both strains, but Crm1 expression and levels of E2f4-Crm1 complex were higher in F344 rats. Marked upregulation of P16(INK4A) was associated with moderate overexpression of HSP90, CDC37, E2F4, and CRM1 in human hepatocellular carcinomas with a better prognosis. In contrast, strong induction of HSP90, CDC37, and E2F4 was paralleled by P16(INK4A) downregulation and high levels of HSP90-CDK4 and CDC37-CDK4 complexes in hepatocellular carcinomas with poorer prognosis. CDC37 downregulation by small interfering RNA inhibited in vitro growth of HepG2 cells. In conclusion, our findings underline the role of Hsp90/Cdc37 and E2f4/Crm1 systems in the acquisition of a susceptible or resistant carcinogenic phenotype. The results also suggest that protection by CDC37 and CRM1 against growth restraint by P16(INK4A) influences the prognosis of human hepatocellular carcinoma.

Purpose: To evaluate the association between Heidelberg Retina Tomograph (HRT) parameters and the Frequency Doubling Technology (FDT) indices in ocular hypertension or primary open angle glaucoma patients., Methods: One eye was randomly chosen from 85 patients with primary open angle glaucoma (abnormal visual field and/or abnormal optic nerve and untreated IOP > 21 mm Hg) and ocular hypertension (normal visual field and normal optic nerve and untreated IOP > 21 mm Hg). All the patients were examined with Humphrey Perimeter program 30-2 full threshold, FDT program C 20-5, and HRT. Findings were analyzed using Pearson's correlation coefficient and linear regression., Results: Within the entire sample of studied subjects we found a statistically significant correlation (Pearson's r) (P < 0.05) between FDT mean deviation and RA, CA, RV, CSM, RNFLt, and RNFLc of HRT, and between FDT pattern standard deviation and RA, CA, VOLC, CV, RV, CSM, RNFLt, and RNFLc of HRT. Linear regression analysis showed that cup shape measure was the most predictive parameter for FDT MD. No correlation was found between HRT parameters and FDT PSD indices in the glaucoma subgroup; in the ocular hypertensive subgroup some correlation was found between FDT PSD and some of the HRT parameters., Conclusion: In the entire group, cup shape measure showed to be the most significant factor correlating functional and morphologic changes. The presence of significant correlations between some HRT parameters and visual field indices in the ocular hypertensive and glaucoma subgroup suggested that HRT parameters and FDT indices could detect glaucoma damage differently.

Preneoplastic and neoplastic hepatocytes undergo c-Myc up-regulation and overgrowth in rats genetically susceptible to hepatocarcinogenesis, but not in resistant rats. Because c-Myc regulates the pRb-E2F pathway, we evaluated cell cycle gene expression in neoplastic nodules and hepatocellular carcinomas (HCCs), induced by initiation/selection (IS) protocols 40 and 70 weeks after diethylnitrosamine treatment, in susceptible Fisher 344 (F344) rats, and resistant Wistar and Brown Norway (BN) rats. No interstrain differences in gene expression occurred in normal liver. Overexpression of c-myc, Cyclins D1, E, and A, and E2F1 genes, at messenger RNA (mRNA) and protein levels, rise in Cyclin D1-CDK4, Cyclin E-CDK2, and E2F1-DP1 complexes, and pRb hyperphosphorylation occurred in nodules and HCCs of F344 rats. Expression of Cdk4, Cdk2, p16(INK4A), and p27(KIP1) did not change. In nodules and/or HCCs of Wistar and BN rats, low or no increases in c-myc, Cyclins D1, E, and A, and E2F1 expression, and Cyclin-CDKs complex formation were associated with p16(INK4A) overexpression and pRb hypophosphorylation. In conclusion, these results suggest deregulation of G1 and S phases in liver lesions of susceptible rats and block of G1-S transition in lesions of resistant strains, which explains their low progression capacity.

Hepatocarcinogenesis sensitivity (Hcs1, 2) and resistance (Hcr1-3) loci have been identified by linkage analysis on rat chromosomes 7 and 1, and 10, 4, and 8, respectively. Cytogenetic studies documented deletions on chromosomes 3 and 6 of neoplastic rat hepatocytes. Hepatocellular carcinomas (HCCs) were produced in F1 hybrid rats between Long-Evans (LE) and Fisher 344 (F344) rats. Scanning of the above chromosomes for loss of heterozygosity (LOH) showed allelic imbalance (AI) at multiple regions on chromosomes 6, 7, and 10q. Detailed deletion mapping of chromosome 10 localized a putative suppressor Hcr1 gene to within a 3.2-cM interval flanked by D10Rat51 and D10Rat121. Two other distinct regions with frequent AIs were found inside the Hcr1 locus, at marker loci including DNaseI and Mrp genes, and in a segment including 4 consecutive markers (D10Rat64, D10Rat182, D10Rat113, D10Rat216). In 40% of HCCs, AI was seen at the p53 locus. AI on chromosome 7 occurred at the Hcs1 locus, where is located c-myc, which is amplified in HCCs, suggesting allelic gain. Most AIs occurred in poorly/moderately differentiated carcinomas, and a few events were seen in well-differentiated tumors on chromosomes 7 and 10. These data suggest that alteration of a cluster of oncosuppressor genes on 10q is important for HCC progression. The existence of AI on segments of rat chromosomes 6, 7, and 10, syntenic to chromosomal segments of human HCCs where chromosomal gains or deletions occur, suggests a commonality of some molecular events in the pathogenesis of HCCs in rats and humans. Our map provides information toward cloning putative oncosuppressor genes associated with this carcinoma.

Molecular mechanisms of basal and D-amphetamine (AMPH)-induced apoptosis were studied in rat liver nodules, 12 (N12) and 30 (N30) weeks after initiation, and in hepatocellular carcinoma (HCC) induced by diethylnitrosamine in rats subjected to resistant hepatocyte model. Basal apoptosis in hematoxylin/eosin- and propidium iodide-stained sections was higher in nodules and HCC than in normal livers. It sharply increased in all tissues 4 hours after AMPH treatment (10 mg/kg), and declined to basal levels at 8 to 12 hours in liver and N12, but remained high up to 18 hours in N30 and HCC. c-myc, Tgf-alpha, p53, and Bcl-X(S) messenger RNA (mRNA) levels were higher, and Bcl-2 mRNA was lower in N12 and/or N30 and HCC than in normal liver. Four hours after AMPH injection, increase in c-myc and decreases in Bcl-2 and Bcl-X(L) mRNAs occurred in all tissues, whereas p53, Bax, and Bcl-X(S) mRNAs increased in N30 and HCC. These changes disappeared in liver and N12 at 18 hours, but persisted in N30 and HCC. c-Myc, P53, Bcl-2, and Bax proteins in normal liver and HCC +/- AMPH showed similar patterns. Tgf-beta1, Tgf-beta-RIII, CD95, and CD95L mRNA levels underwent slight or no changes in any tissue +/- AMPH. Basal Hsp27 expression was high in nodules and HCC, and was stimulated by AMPH in liver and N12, but not in N30 and HCC. These data suggest a role of dysregulation of Bcl-2 family genes and, at least in atypical lesions, of p53 overexpression, in basal and AMPH-induced apoptosis in nodules and HCCs. Hsp27 does not appear to sufficiently protect atypical lesions against apoptosis.

Iron plays an important role in cell growth and metabolism. In preneoplastic liver nodules, a rise in the number of transferrin receptors (Tf-R) is associated with decreased endocytosis of the Fe2-Tf/Tf-R complex. Because nodules are precursors of hepatocellular carcinoma (HCC), the question arises whether changes in iron uptake by nodules persist in HCC. Current work showed up-regulation of Tf messenger RNA (mRNA) production in preneoplastic nodules, 12 to 37 weeks after initiation, and down-regulation in atypical nodules (at 45 and 50 weeks) and HCCs, induced in rats by the "resistant hepatocyte" model. Tf-R gene expression increased in nodules and HCCs. Tf-R numbers increased, without changes in affinity constant, in HCC. Iron uptake was higher in HCC than in normal liver, 5 to 40 minutes after injection of 59Fe2-Tf, with preferential accumulation in cytosol of tumor cells and in microsomes of normal liver. Purification through Percoll gradient of mitochondria plus lysosomes allowed the identification in liver and HCC of an endosomal compartment sequestering injected 125I-Tf. This subfraction was not seen when 59Fe2-Tf was injected into rats, and 59Fe was found in particulate material of both tissues. Liver and HCC exhibited comparable basal activities of plasma membrane NADH oxidase, an enzyme involved in iron uptake and cell growth. Stimulation of this activity by Fe2-Tf was higher in HCC than in normal liver. These results indicate that Tf expression may be a marker of preneoplastic liver progression to malignancy. Differently from nodules, HCC may sequester relatively high iron amounts, necessary for fast growth, both through the endocytic pathway and the reduced form of nicotinamide adenine dinucleotide (NADH) oxidase system.