Assessment of clinically significant portal hypertension by two-dimensional shear wave elastography.

BACKGROUND AND AIMS: To evaluate two-dimensional shear wave elastography (2DSWE) in parallel with transient elastography (TE) for diagnosing clinically significant portal hypertension (CSPH) and high-risk varices (HRV) in patients with chronic liver disease. PATIENTS AND METHODS: Consecutive patients with suspicion of compensated advanced chronic liver disease (cACLD) [liver stiffness measurement (LSM) ≥ 10 kPa by TE, or morphological signs suggestive of cACLD on imaging], with no history of liver decompensation, underwent hepatic venous pressure gradient (HVPG) measurement, transjugular liver biopsy and esophagogastroduodenoscopy, which served as the reference methods for diagnosing CSPH, cACLD and HRV. All patients underwent LSM and spleen stiffness measurements (SSM) by 2DSWE and TE. RESULTS: Seventy-six (76) patients were included (78% men, mean age 62 years, body mass index 28.3 kg/m2 , 36.8% alcoholic, 30.3% non-alcoholic fatty liver disease, 14.5% viral hepatitis). Of them, 80.3%, 69.7%, 52.6% and 22.4% had cACLD, cirrhosis, CSPH and HRV respectively. LSM performed better than SSM in diagnosing CSPH and HRV. For CSPH, AUROCs (0.926 vs. 0.866), optimal cut-offs (20.1 vs. 20.2 kPa) and sensitivity/specificity (80.5%/94.3% vs. 77.5% /86.1%) were comparable for 2DSWE and TE. Ruling-out of CSPH by 2DSWE (LSM at cut-off with ≥90% sensitivity (13.5 kPa) and platelets ≥ 150 x 109 /L) performed comparably to TE, with 1/24 cases falsely classified as negative. For HRV, AUROCs were similar (0.875 2DSWE, 0.851 TE) with similar optimal LSM cut-offs enabling 100% sensitivity and ruling-out HRV. CONCLUSION: Liver stiffness measurement by 2DSWE appears to perform equally well as TE for diagnosing CSPH and ruling-out HRV in compensated chronic liver disease.

Claudins: Beyond Tight Junctions in Human IBD and Murine Models.

Claudins are transmembrane proteins constituting one of three tight junction protein families. In patients with inflammatory bowel disease (IBD), disease activity-dependent changes in expression of certain claudins have been noted, thus making certain claudin family members potential therapy targets. A study was undertaken with the aim of exploring expression of claudins in human disease and two different animal models of IBD: dextrane sulfate sodium-induced colitis and adoptive transfer model of colitis. The expression of sealing claudin-1, claudin-3, claudin-4, and claudin-8, and pore-forming claudin-2 in humans and rodents has been evaluated by immunohistochemistry and quantitative polymerase chain reaction. Claudins were expressed by epithelial and cells of mesodermal origin and were found to be situated at the membrane, within the cytoplasm, or within the nuclei. Claudin expression by human mononuclear cells isolated from lamina propria has been confirmed by Western blot and flow cytometry. The claudin expression pattern in uninflamed and inflamed colon varied between species and murine strains. In IBD and both animal models, diverse alterations in claudin expression by epithelial and inflammatory cells were recorded. Tissue mRNA levels for each studied claudin reflected changes within cell lineage and, at the same time, mirrored the ratio between various cell types. Based on the results of the study, it can be concluded that 1) claudins are not expressed exclusively by epithelial cells, but by certain types of cells of mesodermal origin as well; 2) changes in the claudin mRNA level should be interpreted in the context of overall tissue alterations; and 3) both IBD animal models that were analyzed can be used for investigating claudins as a therapy target, respecting their similarities and differences highlighted in this study.

Prognostic Significance of Lacunarity in Preoperative Biopsy of Colorectal Cancer.

The quantity and quality of preoperative material in colorectal cancer is often limiting factor in determination of risk factors and therapy planning. The most important negative prognostic factors are intravascular and perineural invasion, as well as tumor budding. Usually, the only parameter available in preoperative biopsy is tumor budding. However, the growing body of evidence suggests that cancer differentiation based on the poorly differentiated clusters has better prognostic value. The limiting factor in applying of these new parameters is reproducible, simple, cheap and fast method of their determination. In this paper we investigated the prognostic value of lacunarity, determined in preoperative biopsy. Lacunarity is a measure of spatial heterogeneity (inhomogeneity) in an image. It quantifies how objects fill the space, and enables analysis of gaps distribution, homogeneity of gaps, and presence of structures. It was shown that lacunarity and the total number of buds could be combined in a model which clearly divides colorectal cancer patients in low, medium and high risk subgroups. The paper also points out that the quantitative numerical methods are superior to semiquantitative methods, and that individual methods should be combined using algorithms to obtain a more accurate prediction. Because the study described is designed as a pilot study, verification is needed on a larger sample of patients from independent researchers.

Insulin-like growth factor 2 binding protein 3 expression on endoscopic ultrasound guided fine needle aspiration specimens in pancreatic ductal adenocarcinoma.

BACKGROUND: Despite numerous investigations, we still do not have a specific marker for pancreatic ductal adenocarcinoma. Only guideline-recommended biomarker for pancreatic ductal adenocarcinoma is the CA19-9, but it is also present in other gastrointestinal diseases. IMP3 is a new potential biomarker that is over-expressed in some cancers. The aims of our study were (1) to assess IMP3 in benign pancreatic lesions and pancreatic cancer, and (2) to estimate its concentrations in localized and advanced pancreatic cancer. PATIENTS AND METHODS: Seventy-five patients with solid pancreatic lesions who underwent EUS-FNA were included. Patients were divided into three groups: benign lesions, cancer localized only on the pancreas, and patients with advanced pancreatic cancer (locally advanced or with distal metastases). Immunoreactivity of IMP3 was assessed on cytological smears sampled by endoscopic ultrasound. RESULTS: IMP3 was expressed in 89% of the patients with pancreatic cancer and not in benign lesions. Stronger expression of IMP3 protein and stage of the pancreatic cancer was statistically significant. IMP3 was expressed in all localized cancers and in 85% of patients with advanced pancreatic cancer. In the subgroup with locally advanced cancer, IMP3 was expressed in 88%, and in 83% of patients in the subgroup with distal metastasis (P = 0.007). In the present study, sensitivity was 89%, specificity 100%, with positive predictive value of 100% and negative predictive value of 63%. CONCLUSION: There is a positive correlation between IMP3 expression and TNM stages of the pancreatic cancer. Higher expression of IMP3 on EUS-FNA specimens can suggest poorer prognosis.

Prothrombin 3'end Gene Variants in Patients With Sporadic Colon Adenocarcinoma.

BACKGROUND/AIM: Thrombin plays significant roles in various types of cancer. However, the expression levels of prothrombin, the thrombin precursor, in cancer remain unclear. Variants of the 3'end of the prothrombin gene lead to increased prothrombin expression. This study aimed to analyze prothrombin 3'end gene variants in colon tumor and adjacent normal tissue samples. MATERIALS AND METHODS: The study group consisted of 93 patients suffering from colon adenocarcinoma. The 3'end of the prothrombin gene was analyzed by DNA sequencing. RESULTS: Three variants, all previously associated with increased prothrombin expression were detected. Frequency of the FII 19911G allele was 46.77% and 47.85% in tumor and normal tissue, respectively. For the FII 20210A allele, the detected frequencies were 2.15% and 1.61%, respectively. The frequency of the FII c.1824T allele was 0.54% in both tissues. Four patients showed different genotypes in tumor and normal tissue. CONCLUSION: Prothrombin 3' end gene variants may play a role in colorectal cancer.

Polymorphisms in Survivin (BIRC5 Gene) Are Associated with Age of Onset in Breast Cancer Patients.

Survivin, encoded by BIRC5 gene (baculoviral IAP repeat containing 5), belongs to the family of inhibitors of apoptosis proteins (IAPs). In mammalian cells it participates in the control of mitosis, apoptosis regulation, and cellular stress response. Its expression is increased in almost all types of cancers. The aim of this study was to investigate the role of BIRC5 polymorphisms in breast cancer (BC) and to connect survivin expression with various clinicopathological characteristics of BC patients. Blood and archival tumour tissue samples were collected from 26 BC patients from Croatia. Survivin expression was determined immunohistochemically. BIRC5 promoter, coding region, and 3'UTR were genotyped. DNA from 74 healthy women was used as control. BIRC5 polymorphisms and survivin expression were tested against age of onset, histological grade, tumour type and size, lymph node status, oestrogen, progesterone, Her2, and Ki67 status. Numbers of samples with weak, moderate, and strong survivin expression were 9 (33.3%), 11 (40.7%), and 7 (25.9%), respectively. Most patients had nuclear survivin staining (92.6%). High survivin expression was significantly associated with negative oestrogen receptor status (p=0.007) and positive Ki67 expression (p=0.032). Ki67 expression was also positively correlated with histological grade (p=0.0009). Fourteen polymorphisms were found in BC samples, located mostly in promoter and 3'UTR of BIRC5. There was no significant difference in the distribution of polymorphisms between BC and control samples. Among clinicopathological characteristics of BC patients, alleles of five BIRC5 polymorphisms were associated with younger age of onset: c.-644T>C (55.8 years [y] vs. 48.1 y; p=0.006), c.-241C>T (54.2 y vs. 45.0; p=0.029), c.9809T>C (55.8 y vs. 48.1 y; p=0.006), c.-1547C>T (58.3 y vs. 50.9 y; p=0.011), and c.9386T>C (50.8 y vs. 59.5 y; p=0.004). To assess the significance of BIRC5 polymorphisms and survivin expression as predictive and prognostic biomarkers for BC further research with a larger sample size is needed.

Targeting Tumor Markers with Antisense Peptides: An Example of Human Prostate Specific Antigen.

The purpose of this paper was to outline the development of short peptide targeting of the human prostate specific antigen (hPSA), and to evaluate its effectiveness in staining PSA in human prostate cancer tissue. The targeting of the hPSA antigen by means of antisense peptide AVRDKVG was designed according to a three-step method involving: 1. The selection of the molecular target (hPSA epitope), 2. the modeling of an antisense peptide (paratope) based on the epitope sequence, and 3. the spectroscopic evaluation of sense-antisense peptide binding. We then modified standard hPSA immunohistochemical staining practice by using a biotinylated antisense peptide instead of the standard monoclonal antibody and compared the results of both procedures. Immunochemical testing on human tissue showed the applicability of the antisense peptide technology to human molecular targets. This methodology represents a new approach to deriving peptide ligands and potential lead compounds for the development of novel diagnostic substances, biopharmaceuticals and vaccines.

Albi Score as a Predictor of Survival in Patients with Compensated Cirrhosis Resected for Hepatocellular Carcinoma: Exploratory Evaluation in Relationship to Palbi and Meld Liver Function Scores.

The aim of the study was to explore predictive value of the ALBI, PALBI and MELD scores on survival in patients resected for hepatocellular carcinoma with compensated liver cirrhosis and no macrovascular infiltration. In this retrospective study, longitudinal survival analysis was performed. We analyzed patient/tumor characteristics and MELD, ALBI and PALBI scores as liver function tests for predicting survival outcome. Survival was analyzed from the date of liver resection until death, liver transplantation, or end of follow-up. Patients were stratified for age, cirrhosis etiology, presence of esophageal varices, hepatocellular carcinoma stage, microvascular invasion, histologic differentiation, and resection margins. We identified 38 patients (alcoholic cirrhosis in 84.2% of patients) resected over an 8-year period. Median preoperative MELD score was 8, ALBI score -2.63, and PALBI score -2.38. During the follow-up period, 24 patients died. Estimated median survival time was 36 months. Microvascular invasion was observed in 33 patients. Higher ALBI score was associated with 23.1% higher relative risk of death. PALBI score was associated with 12.1% higher relative risk of death, whereas MELD score was not associated with the risk of death. In conclusion, ALBI score demonstrated significant predictive capabilities for survival in patients with compensated cirrhosis resected for hepatocellular carcinoma.

Analysis of MSH2 Loss of Heterozygosity, Expression, and IVS10+12G>A Polymorphism in Sporadic Colon Cancer.

BACKGROUND: mutS homolog 2 (MSH2) deficiency may be involved in the development of microsatellite instability found in certain sporadic colorectal tumors. In addition to mutations or loss of heterozygosity resulting in complete loss of MSH2 function, polymorphisms affecting MSH2 expression have been also identified. Therefore, the aim of this study was to examine MSH2 status in sporadic colon cancer. MATERIALS AND METHODS: MSH2 status was examined at the DNA, RNA and protein levels through loss of heterozygosity (LOH) analysis, quantitative real-time PCR and immunohistochemistry. MSH2 IVS10+12A>G polymorphism was examined by real-time single nucleotide polymorphism genotyping. RESULTS: MSH2 LOH was more frequent in tumors larger than 5 cm (p=0.032), mRNA expression was also significantly lower and the same expression pattern was present in the corresponding normal mucosa of the same patient (p=0.013 and p=0.008, respectively). No association was found between IVS10+12A>G polymorphism and susceptibility to sporadic colon cancer. CONCLUSION: Altered MSH2 expression detected in sporadic colon tumors pointing to its role in colorectal tumorigenesis without a hereditary component.

Comparison of hepatoprotective effect from ischemia-reperfusion injury of remote ischemic preconditioning of the liver vs local ischemic preconditioning of the liver during human liver resections.

AIM: To compare and evaluate the hepatoprotective effect of remote ischemic preconditioning (RIPC) with local ischemic preconditioning (LIPC) of the liver during human liver resections. METHODS: A prospective, single-centre, randomised control trial was conducted in the Clinical Hospital "***" from April 2017 to January 2018. A total of 60 patients, who underwent liver resection due to colorectal cancer liver metastasis, were randomised to one of three study arms: 1) a RIPC group, 2) an LIPC group and 3) a control group (CG) in which no ischemic preconditioning was done before liver resection. The hepatoprotective effect was evaluated by comparing serum transaminase levels, bilirubin levels, albumin, and protein levels, coagulograms and through pathohistological analysis. The trial was registered on ClinicalTrials.gov (NCT****). RESULTS: Significant differences were found in serum levels of liver transaminases and bilirubin levels between thegroups, the highest level in the CG and the lowest level in the LIPC group. Levels of cholinesterase were also significantly higher in the LIPC group. Pathohistological findings graded by the Rodriguez score showed favourable changes in the LIPC and RIPC groups versus the CG. CONCLUSION: Strong evidence supports the hepatoprotective effect of RIPC and LIPC preconditioning from an ischemia-reperfusion injury of the liver. Better synthetic liver function preservation in these two groups supports this conclusion.

High absolute basophil count is a powerful independent predictor of inferior overall survival in patients with primary myelofibrosis.

OBJECTIVES: To investigate the clinical and prognostic significance of absolute basophil count (ABC) in patients with primary myelofibrosis (PMF). METHODS: We retrospectively investigated 58 patients with PMF treated in our institution in the period from 2006 to 2017. ABC was obtained in addition to other hematological and clinical parameters. Patients were separated into high and low ABC groups using the Receiver operating characteristic curve analysis. RESULTS: ABC was higher in PMF patients than in healthy controls (P < 0.001). Patients with high ABC had higher white blood cells (P < 0.001), higher red cell distribution width (P = 0.035), higher lactate dehydrogenase (P < 0.001), more frequently had circulatory blasts (P < 0.001), constitutional symptoms (P = 0.030) and massive splenomegaly (P = 0.014). ABC was also positively correlated with absolute monocyte count (AMC) (P < 0.001) and other components of differential blood count. There was no difference in ABC regarding driver mutations or degree of bone marrow fibrosis. Univariately, high ABC was significantly associated with inferior overall survival (hazard ratio (HR) 4.79, P < 0.001). This effect remained statistically significant (HR 4.27, P = 0.009) in a multivariate Cox regression model adjusted for age, gender, Dynamic International Prognostic Scoring System (HR 2.6, P = 0.001) and AMC (HR 8.45, P = 0.002). DISCUSSION: High ABC reflects higher disease activity and stronger proliferative potential of disease. ABC and AMC independently predict survival and therefore seem to reflect different underlying pathophysiologic processes. Hence, both have a potential for improvement of current prognostic scores. CONCLUSION: Basophils represent a part of malignant clone in PMF and are associated with unfavorable disease features and poor prognosis which is independent of currently established prognostic scoring system and monocytosis.

Liver elastography malignancy prediction score for noninvasive characterization of focal liver lesions.

BACKGROUND & AIMS: To analyse elastographic characteristics of focal liver lesions (FLL)s and diagnostic performance of real-time two-dimensional shear-wave elastography (RT-2D-SWE) in order to differentiate benign and malignant FLLs. METHODS: Consecutive patients diagnosed with FLL by abdominal ultrasound (US) underwent RT-2D-SWE of FLL and non-infiltrated liver by intercostal approach over the right liver lobe. The nature of FLL was determined by diagnostic work-up, including at least one contrast-enhanced imaging modality (MDCT/MRI), check-up of target organs when metastatic disease was suspected and FLL biopsy in inconclusive cases. RESULTS: We analysed 196 patients (median age 60 [range 50-68], 50.5% males) with 259 FLLs (57 hepatocellular carcinomas, 17 cholangiocarcinomas, 94 metastases, 71 haemangiomas, 20 focal nodular hyperplasia) of which 70 (27%) were in cirrhotic liver. Malignant lesions were stiffer (P < .001) with higher variability in intralesional stiffness (P = .001). The best performing cut-off of lesion stiffness was 22.3 kPa (sensitivity 83%; specificity 86%; positive predictive value [PPV] 91.5%; negative predictive value [NPV] 73%) for malignancy. Lesion stiffness <14 kPa had NPV of 96%, while values >32.5 kPa had PPV of 96% for malignancy. Lesion stiffness, lesion/liver stiffness ratio and lesion stiffness variability significantly predicted malignancy in stepwise logistic regression (P < .05), and were used to construct a new Liver Elastography Malignancy Prediction (LEMP) score with accuracy of 96.1% in validation cohort (online calculator available at http://bit.do/lemps). CONCLUSION: The comprehensive approach demonstrated in this study enables correct differentiation of benign and malignant FLL in 96% of patients by using RT-2D-SWE.

Offset-sparsity decomposition for automated enhancement of color microscopic image of stained specimen in histopathology.

We propose an offset-sparsity decomposition method for the enhancement of a color microscopic image of a stained specimen. The method decomposes vectorized spectral images into offset terms and sparse terms. A sparse term represents an enhanced image, and an offset term represents a "shadow." The related optimization problem is solved by computational improvement of the accelerated proximal gradient method used initially to solve the related rank-sparsity decomposition problem. Removal of an image-adapted color offset yields an enhanced image with improved colorimetric differences among the histological structures. This is verified by a no-reference colorfulness measure estimated from 35 specimens of the human liver, 1 specimen of the mouse liver stained with hematoxylin and eosin, 6 specimens of the mouse liver stained with Sudan III, and 3 specimens of the human liver stained with the anti-CD34 monoclonal antibody. The colorimetric difference improves on average by 43.86% with a 99% confidence interval (CI) of [35.35%, 51.62%]. Furthermore, according to the mean opinion score, estimated on the basis of the evaluations of five pathologists, images enhanced by the proposed method exhibit an average quality improvement of 16.60% with a 99% CI of [10.46%, 22.73%].

Unsupervised segmentation of low-contrast multichannel images: discrimination of tissue components in microscopic images of unstained specimens.

Low-contrast images, such as color microscopic images of unstained histological specimens, are composed of objects with highly correlated spectral profiles. Such images are very hard to segment. Here, we present a method that nonlinearly maps low-contrast color image into an image with an increased number of non-physical channels and a decreased correlation between spectral profiles. The method is a proof-of-concept validated on the unsupervised segmentation of color images of unstained specimens, in which case the tissue components appear colorless when viewed under the light microscope. Specimens of human hepatocellular carcinoma, human liver with metastasis from colon and gastric cancer and mouse fatty liver were used for validation. The average correlation between the spectral profiles of the tissue components was greater than 0.9985, and the worst case correlation was greater than 0.9997. The proposed method can potentially be applied to the segmentation of low-contrast multichannel images with high spatial resolution that arise in other imaging modalities.

Mononucleotide repeats in the SMAD4 gene promoter in colon carcinoma tissue of Croatian patients.

This study was aimed at the analysis of mononucleotide repeats -462T(15) and -4T(12) in the SMAD4 gene promoter in sporadic colon adenocarcinoma tissue of Croatian patients. The analysis has included 60 pairs of samples of colon tumor and adjacent normal tissue. The number of thymidines in the tracts -462T(15) and -4T(12) of the SMAD4 gene promoter was determined by PCR with fluorescently labeled primers followed by the analysis of obtained DNA fragments by capillary electrophoresis. In the normal colon tissue two haplotypes were present: -462T(15)/-4T(12) in 51 patients (85%) and -462T(16)/-4T(12) in 9 patients (15%). Among the cases with haplotype -462T(15)/-4T(12) detected in normal colon tissue, in 5 cases (8%) malignant tissue displayed different haplotypes: 462T(10)/-4T(10), -462T(12)/-4T(12), 462T(13)/-4T(11), -462T(14)/-4T(10) and -462T(15)/-4T(11). Haplotype -462T(14)/-4T(10) was previously found to be associated with significantly decreased SMAD4 gene promoter activity in comparison to the wild type, while the other detected haplotypes remain to be functionally characterized. This study has shown that functionally relevant somatic alterations of the SMAD4 gene promoter are found in some colon cancer tumors. Although not as frequent in colon as in pancreatic cancer, they may be of significance for certain cases and their role in colon tumorigenesis should be investigated further.

Modulation of γ2-MSH hepatoprotection by antisense peptides and melanocortin subtype 3 and 4 receptor antagonists.

Melanocortins, i.e., melanocyte stimulating hormones (MSH) are peptides with strong antiinflammatory effects. The most investigated aspects of γ2-MSH are related to cardiovascular effects and natriuresis, with limited research available about its anti-inflammatory and cytoprotective effects. The aims of this study were: 1) to examine the effects of γ2-MSH and its derivative [D-Trp(8)]-γ2-MSH on the acetaminophen model of liver damage in CBA mice; 2) to evaluate the modulation of γ2-MSH hepatoprotection by melanocortin subtypes 3 and 4 receptor antagonists SHU 9119 and HS 024; 3) to define the importance of central MSH pharmacophore region (HFRW) by using antisense peptides LVKAT and VKAT. In this study, specific antagonists and antisense peptides were used to target central pharmacophore region of γ2-MSH and [D-Trp(8)]-γ2-MSH, enabling the evaluation of hepatoprotection from the standpoint of the receptor and pharmacophore blockade. The criteria for monitoring the effects of the hormones on the liver damage were alanine transaminase, aspartate transaminase activities (U/L), and pathohistological scoring of liver necrosis (scale 0-5). γ2-MSH (0.24 mg/kg) indicated hepatoprotective effects in comparison to control (p < 0.001). In contrast, [D-Trp(8)]-γ2-MSH did not show any hepatoprotective effects. Application of antagonists SHU 9119 and HS 024, and antisense peptides LVKAT and VKAT, also did not show any hepatoprotective effects. In fact, when combined with γ2-MSH, it annulled its hepatoprotective effect. The results provide evidence for hepatoprotective and antiinflammatory effects of the γ2-MSH in the liver.

Female headstart in resistance to hyperoxia-induced oxidative stress in mice.

Increased oxygen concentration (hyperoxia) induces oxidative damage of tissues and organs. Oxygen toxicity in hyperoxia is controlled by factors such as sex, age, tissue, strain and hormones. In most species females show lower incidence of some age-related pathologies linked with oxidative stress, which has been attributed to a beneficial effect of ovarian hormones. In this study we found that hyperoxia induced hepatic oxidative damage exclusively in male CBA/H mice, followed by their decreased survival. Histopathological examination revealed that the observed differences in survival were not the consequence of acute lung injury induced by hyperoxia. Next, we observed that an increased Sirt1 protein level in hyperoxia-exposed female CBA/H mice correlated with their lower PPAR-γ and higher eNOS and Sod2 protein levels. In males, higher PPAR-γ and lower Sod2 protein levels were associated with unchanged Sirt1 expression. Although these results are of a correlative nature only, they clearly show that females show better survival, increased resistance to hyperoxia and have generally more efficient defense systems, which suggests that their headstart in resistance to hyperoxia could be a consequence of the beneficial effect of ovarian hormones.

Hepatoprotective effects of Met-enkephalin on acetaminophen-induced liver lesions in male CBA mice.

Recent histopathological investigations in patients with hepatitis suggested possible involvement of Met-enkephalin and its receptors in the pathophysiology of hepatitis. Consequently, we evaluated the potential hepatoprotective effects of this endogenous opioid pentapeptide in the experimental model of acetaminophen induced hepatotoxicity in male CBA mice. Met-enkephalin exhibited strong hepatoprotective effects in a dose of 7.5 mg/kg, which corresponds to the protective dose reported for several different animal disease models. In this group plasma alanine aminotransferase and aspartate aminotransferase enzyme activities, as well as liver necrosis score were significantly reduced in comparison to control animals treated with physiological saline (p>0.01). The specificity of the peptide hepatoprotection was investigated from the standpoint of the receptor and peptide blockade. It was concluded that Met-enkephalin effects on the liver were mediated via δ and ζ opioid receptors. Genotoxic testing of Met-enkephalin confirmed the safety of the peptide.