Centrilobular Zone Research Articles

Morphological changes in the liver parenchyma in rats under subacute acrylamide intoxication and the possibility of their preventive treatment

Introduction. Acrylamide, used in industry, is also formed in foods during high temperature processing. Acrylamide has a toxic effect on the nervous, reproductive, and other systems of the body, due to carcinogenic and genotoxic properties. Considering sufficiently serious threat of acrylamide to human health, it is relevant to carry out further fundamental research to study the mechanism of its impact on the body and the possibility of treatment of lesions caused by it.
 The aims of this study is to assess morphological changes in the liver of experimental animals under acute acrylamide intoxication and study the possibility of their preventive treatment with complex compounds based on oxymethyluracil.
 Materials and methods. Experimental studies on the subacute effects of acrylamide at a dose of 20 mg/kg on the histological structure of the liver were carried out on outbred female rats. The efficiency of preventive administration of complex compounds of oxymethyluracil with ascorbic acid, with sodium succinate and acetylcysteine ​​was also studied.
 Results. Exposure to acrylamide for 28 days at a dose of 20 mg/kg of body weight in experimental rats, has been established to cause minor morphological changes in the structure of the liver: only focal cellular infiltrates were found in the centrilobular zones. In groups of rats that received preventive treatment with complex compounds based on oxymethyluracil, no formation of infiltrates and other pathological changes was observed.
 Limitations. Limitations of the study are related to implementation of experimental studies of the pathomorphological changes in the liver of experimental animals using only one dose of acrylamide - 20 mg per kg of body weight during one limited period (28 days).
 Conclusion. Exposure to acrylamide for 28 days at a dose of 20 mg/kg of body weight causes minor morphological changes in the structure of the liver in experimental rats. For the first time, the effectiveness of prophylactic correction of the toxic effect of acrylamide with complex compounds of oxymethyluracil with ascorbic acid, sodium succinate and acetylcysteine, which showed a certain protective effect on the liver, was studied.

Targeting IKKβ Activity to Limit Sterile Inflammation in Acetaminophen-Induced Hepatotoxicity in Mice.

The kinase activity of inhibitory κB kinase β (IKKβ) acts as a signal transducer in the activating pathway of nuclear factor-κB (NF-κB), a master regulator of inflammation and cell death in the development of numerous hepatocellular injuries. However, the importance of IKKβ activity on acetaminophen (APAP)-induced hepatotoxicity remains to be defined. Here, a derivative of caffeic acid benzylamide (CABA) inhibited the kinase activity of IKKβ, as did IMD-0354 and sulfasalazine which show therapeutic efficacy against inflammatory diseases through a common mechanism: inhibiting IKKβ activity. To understand the importance of IKKβ activity in sterile inflammation during hepatotoxicity, C57BL/6 mice were treated with CABA, IMD-0354, or sulfasalazine after APAP overdose. These small-molecule inhibitors of IKKβ activity protected the APAP-challenged mice from necrotic injury around the centrilobular zone in the liver, and rescued the mice from hepatic damage-associated lethality. From a molecular perspective, IKKβ inhibitors directly interrupted sterile inflammation in the Kupffer cells of APAP-challenged mice, such as damage-associated molecular pattern (DAMP)-induced activation of NF-κB activity via IKKβ, and NF-κB-regulated expression of cytokines and chemokines. However, CABA did not affect the upstream pathogenic events, including oxidative stress with glutathione depletion in hepatocytes after APAP overdose. N-acetyl cysteine (NAC), the only FDA-approved antidote against APAP overdose, replenishes cellular levels of glutathione, but its limited efficacy is concerning in late-presenting patients who have already undergone oxidative stress in the liver. Taken together, we propose a novel hypothesis that chemical inhibition of IKKβ activity in sterile inflammation could mitigate APAP-induced hepatotoxicity in mice, and have the potential to complement NAC treatment in APAP overdoses.

High-fat diet-induced nonalcoholic steatohepatitis is accelerated by low carnitine and impaired glucose tolerance in novel murine models

AbstractCarnitine deficiency and impaired glucose tolerance (IGT) exacerbate liver steatosis. Given the current lack of ideal murine nonalcoholic steatohepatitis (NASH) models, we investigated new NASH models using jvs/+ mice with low carnitine and wild-type mice with low-dose alloxan-induced IGT. The jvs/+ and wild-type mice were divided into jvs/+ mice fed a high-fat diet (HFD) from 3 weeks of age (HF hetero group), wild-type mice with low-dose alloxan treatment fed HFD (AL + HF wild group), wild-type mice fed HFD (HF wild group), and two types of mice fed a normal diet—jvs/+ and wild-type (intact group). All mice were sacrificed at 20 or 40 weeks of age. All male HFD-fed mice showed obesity, IGT, high blood insulin levels, homeostatic model assessment of insulin resistance (HOMA-IR), high liver enzyme levels, and high cholesterol levels. The degree of IGT was the worst in the AL + HF wild group, and blood insulin levels and HOMA-IR score were remarkably increased from 20 to 40 weeks of age. Almost all HFD-fed mice showed steatosis, fibrosis, and lobular inflammation in the centrilobular zone. These changes were accompanied by hepatocyte ballooning and were enhanced at 40 weeks of age. Furthermore, the incidence rate of nodular hyperplasia and adenoma in both the HF hetero and AL + HF wild groups was nearly 30%. We successfully established two novel murine models of NASH using male jvs/+ mice with low carnitine and male wild-type mice with IGT that eventually developed obesity, fatty liver, insulin resistance, liver fibrosis, and tumorigenesis. These results suggest that low carnitine levels and early-stage induction of IGT are important factors in the progression of NASH to tumorigenesis, similar to human NASH.

A review of hepatic fibrosis-associated histopathology in aged cadavers.

This article reviews hepatic fibrosis-associated histopathology of aged cadavers (mean age 82 years). A study of 68 livers identified steatosis in 35.5%, central vein fibrosis in 49.2%, perisinusoidal fibrosis in 63.2%, portal tract fibrosis in 47.7%, septa formation in 44.1%, bridging fibrosis in 30.8%, and cirrhosis in 4.4% of the samples as well as one hepatocellular carcinoma and six metastatic tumors. Other studies have revealed that collagens I, III, IV, V, and VI and fibronectin constitute the matrices of fibrous central veins, perisinusoidal space, portal tracts, and septa. Elastin is rich in portal tracts and fibrous septa but absent from the perisinusoidal space. Hepatic stellate cells are ubiquitous in the liver parenchyma while myofibroblasts localize in fibrotic foci. Factor VIII-related antigen expression signals sinusoidal to systemic vascular endothelium transformation while collagen IV and laminin codistribution indicates formation of perisinusoidal membranes. Their coincidence reflects focalized capillarization of sinusoids in the aged liver. In response to fibrogenesis, hepatic progenitor cells residing in the canal of Hering in the periportal parenchyma undergo expansion and migration deep into the lobule. Concomitantly, intermediate hepatocyte-like cells increase in advanced fibrosis stages, which is possibly related to hepatic regeneration. Metabolic zonation of glutamine synthetase expands from the perivenous to non-perivenous parenchyma in fibrosis progression but its expression is lost in cirrhosis, while cytochrome P-4502E1 expression is maintained in centrilobular and midlobular zones in fibrosis progression and expressed in cirrhosis. Hence, cadaveric livers provide a platform for further investigation of hepatic histopathologies associated with the aging liver.

Identification and Spatial Visualization of Dysregulated Bile Acid Metabolism in High-Fat Diet-Fed Mice by Mass Spectral Imaging.

Changes in overall bile acid (BA) levels and specific BA metabolites are involved in metabolic diseases, gastrointestinal, and liver cancer. BAs have become established as important signaling molecules that enable fine-tuned inter-tissue communication within the enterohepatic circulation. The liver, BAs site of production, displayed physiological and functional zonal differences in the periportal zone versus the centrilobular zone. In addition, BA metabolism shows regional differences in the intestinal tract. However, there is no available method to detect the spatial distribution and molecular profiling of BAs within the enterohepatic circulation. Herein, we demonstrated the application in mass spectrometry imaging (MSI) with a high spatial resolution (3 μm) plus mass accuracy matrix-assisted laser desorption ionization (MALDI) to imaging BAs and N-1-naphthylphthalamic acid (NPA). Our results could clearly determine the zonation patterns and regional difference characteristics of BAs on mouse liver, ileum, and colon tissue sections, and the relative content of BAs based on NPA could also be ascertained. In conclusion, our method promoted the accessibility of spatial localization and quantitative study of BAs on gastrointestinal tissue sections and demonstrated that MALDI-MSI was a valuable tool to investigate and locate several BA molecules in different tissue types leading to a better understanding of the role of BAs behind the gastrointestinal diseases.

ANTIOXIDANT EFFECT OF SPINACH EXTRACT IN LIVER FIBROGENESIS ASSOCIATED TO ACTIVATION OF NRF2/HO-1 IN HYPERGLYCEMIC RATS

In chronic hyperglycemia, increased oxidative stress plays an essential role in liver fibrogenesis. The spinach phytochemicals are shown to possess chemopreventive properties in this condition. Erythroid nuclear transcription factor 2 (Nrf2) is a transcriptional regulator expressing cytoprotective genes such as heme oxygenase-1 (ho-1). HO-1 is an enzyme present in the liver with antioxidant, anti-inflammatory, and anti-apoptotic capacities. However, the beneficial effect of spinach associated with endogenous activation of the Nrf2/HO-1 pathway is unknown. Objective: To evaluate the antioxidant effect of the spinach extract on injury hepatic associated with the Nrf2/HO-1 pathway in hyperglycemic rats. The study was approved by the ethics committee from the ENCB-IPN (CONBIOETICA/09/CEI/002/20190327). We used hyperglycemic Wistar rats induced with 60 mg / kg of streptozotocin (HG; n = 7); intragastrical-treated HG with 400mg / kg of spinach methanolic extract (HG-EME; n = 7) and normoglycemic rats (NG; n = 7). Histological sections were obtained at 12 weeks and evaluated by Sirius red staining and immunohistochemistry (Nrf2, HO-1). The oxidative damage by lipid peroxidation was determined by the tissues' malonaldehyde formation (MDA) levels. The H-score system quantified the percentage of nuclear staining of Nrf2 and the intensities of Sirius red and HO-1. Statistical analysis: The data were analyzed by one-way ANOVA with the Tukey test using the GraphPad Prism program (Version 5.0). Kruskal-Wallis and Dunn's test established the MDA training levels. Statistical significance was considered with p <0.05. Collagen fibers were formed mainly in the centrilobular zone. The percentage of collagen fibers staining in the HG group was 50 ± 10, compared to HG-EME (30 ± 10 p <0.05). In NG, the percentage of fibers was 20 ± 10. The nuclear Nrf2 (nNrf2) and cytoplasmic HO-1 staining were localized in the hepatocytes of region 3 Rappaport. The percentage of nNrf2 in HG was 30 ± 10, compared to HG-EME (70 ± 10 p <0.01). In NG, the staining percentage for nNrf2 was 15 ± 5. For HO-1 staining, the values in the HG-EME group were higher than in the HG group (p <0.01). Contrary, MDA levels in HG-EME decreased significantly compared to HG (p <0.05). Treatment with EME in hyperglycemic rats showed decreased liver damage generated by oxidative stress, associated with endogenous activation of the Nrf2/HO-1 pathway. The evaluation of liver biochemical tests can demonstrate a beneficial effect of EME. This work was supported by federal resources (HJM0713/19-I) and partially by INCICH. The authors declare that there is no conflict of interest.

Тканевый и ультраструктурный анализ печени крыс препубертатного возраста при субтоксическом воздействии кадмия и свинца

Using light and electron microscopy, the effect of intoxication with cadmium sulfate (Cd) and lead acetate (Pb) for 3 weeks on the architectonics of the liver parenchyma and ultrastructural features of hepatocytes of 40 prepubertal male Wistar rats was studied, taking into account the model of the classical hepatic lobule. Animals were divided into 4 groups: 1st – control; 2nd – cadmium sulfate solution at a dose of 0.5 mg/kg; 3rd – lead acetate solution 10 mg/kg; 4th – a combination of these solutions. According to stereological analysis of semi-thin sections, statistically significant indicators of periportal zones reflect an increase in the volume of hepatocytes when exposed to Cd (2nd and 4th groups). In all groups, with the exception of the 3rd one (Pb), an increase in the parenchymal compartment in the porto-central gradient was revealed, indicating the formation of hepatocellular ballooning. In all experimental groups, for the periportal zone of the hepatic lobule, a characteristic feature of the ultrastructure of hepatocytes was a statistically significant decrease in the volumetric density of the endoplasmic reticulum; for the centrilobular zone – of mitochondria; for the perivenular zone – mitochondria and nuclei of hepatocytes. The formation of autophagosomes with signs of predominantly mitochondrial utilization attracted attention in groups exposed to Pb. Pathological changes are compensated by increased transsinusoidal metabolism and complexes of biosynthesis organelles.

S2668 Sorafenib-Induced Autoimmune Hepatitis: A Case Report,

Introduction: Drug-induced liver injury (DILI) with autoimmune hepatitis-like features (DIAIH) is a specific phenotype of DILI with clinical features indistinguishable from de-novo autoimmune hepatitis (AIH). We describe a case of DIAIH due to Sorafenib. Case Description/Methods: A 61-year-old man and teetotaller presented with 1-week history of jaundice and malaise. He was on Atorvastatin 40mg daily for 4 years for hyperlipidaemia. Sorafenib was recently initiated six weeks ago for recurrent sarcoma of the left thigh. Clinical examination was unremarkable apart from scleral icterus. Liver function test (LFT) showed severe hepatocellular (HC) injury (bilirubin 4.56 mg/dL, alkaline phosphatase 190 U/L, alanine transaminase 1004 U/L, aspartate transaminase 790 U/L peak INR 1.53). Viral hepatitis screening, AIH specific antibodies and abdominal imaging were unremarkable. Serum IgG was raised (18.6 g/L). Liver biopsy was supportive of DILI and AIH histological features were also noted (Fig.1a-f). Simplified AIH score was 6. Roussel Uclaf Causality Assessment Model (RUCAM) score was 9 for Sorafenib and 6 for Atorvastatin. Diagnosis of Sorafenib-induced AIH was made. His LFT improved spontaneously with normalization of LFT eight weeks after stopping Sorafenib. Discussion: Sorafenib often causes idiosyncratic DILI with mild elevation of transaminases. However, it rarely causes severe DILI and has no known association with DIAIH, unlike Atorvastatin and immune checkpoint inhibitors (e.g Nivolumab and Atezolizumab). To our knowledge, this is the first case report of DIAIH due to Sorafenib. In patients with severe DILI or DILI with probable AIH based on AIH scoring, liver biopsy should be considered for evaluation. However, differentiating DIAIH from de-novo AIH is often challenging as both present with HC injury, may have positive AIH specific antibodies, raised serum IgG and share similar histological findings. The key distinguishing features are the history of drug exposure within a time frame and no requirement for long term immunosuppressant. DIAIH may resolve spontaneously after stopping culprit drug. Corticosteroids should be initiated in cases without improvement upon drug cessation. In DIAIH requiring corticosteroids, relapse is rare after stopping corticosteroids and long-term immunosuppression is rarely needed. In conclusion, DIAIH should be considered in patients who present with severe HC DILI. Differentiating DIAIH from AIH is crucial as the former rarely requires long term immunosuppression.Figure 1.: Liver biopsy specimen [a] H&E 200X: A portal tract, zone 1, showing moderate chronic inflammation with interface damage and rare bile droplets. [b] H&E 200X: Lobulitis characterized by aggregates of plasma cells, swollen hepatocytes and rosetting within liver parenchyma zone 2. [c] H&E 200X: Centrilobular region zone 3 showing hepatocyte drop-out, aggregates of plasma cells, rosetting and bile pigment. Reticulin [d, 40X] and Sirius Red [e, 40X] special stains show collapse with mild early young fibrosis without elastic fibers demonstrated on Orcein [f, 40X], consistent with subacute injury. Overall, the appearances are supportive of subacute DILI in association with features suggestive of AIH (i.e. interface damage, plasma cell aggregates and resetting).

P-47 HISTOPATHOLOGICAL ANALYSIS OF 10-YEAR PROTOCOL LIVER ALLOGRAFT BIOPSY OF ASYMPTOMATIC PEDIATRIC RECIPIENTS FROM A SINGLE BRAZILIAN CENTER

Liver transplant (LT) is the main therapeutic procedure for irreversible liver failure. Protocol liver biopsies during post-transplant follow-up in pediatric recipients with stable graft function have been done in order to identify structural changes. A more specific score, LAFSc, to analyze fibrosis and inflammation on liver allografts has been validated recently. To describe the prevalence of histological liver abnormalities in asymptomatic pediatric patients after ten years following LT from a Brazilian single center comparing two different scoring methods. Cross-sectional study of analysis of protocol percutaneous liver biopsies performed in patients who underwent liver transplant before the age of 18 years, of both genders, asymptomatic, using Tacrolimus for immunosuppression and at least 10-years post-liver transplant. From 97 recipients, 21 met inclusion criteria and had stable liver graft function. A single experienced pathologist assessed histopathological features comparing METAVIR scoring system and LAFSc. Mean follow-up was 12.8 (+/- 2.1) years after post-LT. TAC mean daily dose during biopsy time was 4mg (+/- 1.7) with mean serum level of 4.04. METAVIR scoring system pointed 10 recipients with any degree of fibrosis and 13 with inflammation findings in protocol biopsies. However, using LAFSc scoring system, 13 patients scored for abnormal findings in any of the three zones analysed, although 8 of these had total score of mild fibrosis (1-3). Of the three zones analysed in LAFSc, the highest scores were found in portal space. Centrilobular vein zone was the most affected, documented in 11 recipients. We observed a higher prevalence of abnormal findings using the new allograft fibrosis scoring system – LAFSc. It also showed more specifically the degree and location of fibrosis within hepatic lobule compared to METAVIR. Compared to other LT reports, we observed lower rates of chronic allograft hepatitis and fibrosis at 10 years.

ПАТОМОРФОГЕНЕЗ СТЕАТОЗА ПЕЧЕНИ У ПАЦИЕНТОВ С ОПИОИДНОЙ ЗАВИСИМОСТЬЮ

Background. One of the significant factors in the progression of fibrotic changes in the liver is hepatocyte steatosis, that persists in drug addicted patients even after the elimination of the hepatitis C virus and cessation of drug use. Analysis of the pathomorphogenesis of hepatic steatosis in opioid dependence (OZ) will make it possible to assess the factors that affect ultrastructural changes in hepatocytes and the processes of lipid granule (LH) degradation. Objective. Assessment of ultrastructural changes in LH in the liver tissue of patients with OZ. Material and methods. Histological preparations of liver tissue from 20 patients with OZ aged 21 to 40 years (18 men and 2 women) with different duration of OZ and opioid tolerance. Results. There was established the following dependence of ultrastructural changes in the liver in patients with different duration of OZ and opioid tolerance. The most pronounced changes were noted in the group of patients with prolonged (more than 6 years) opioid intoxication and high tolerance to opioids in the liver tissue, in whom, along with severe steatosis, there were more significant violations of the mechanisms of LH degradation, destruction of cristae in mitochondria, a decrease in the number of lipophagosomes and PH with signs of superficial degradation than in the group of patients with OZ duration up to 6 years, as well as with moderate and high opioid tolerance. Conclusions. Ultrastructural changes in hepatocytes in the form of progression of steatosis in the centrilobular and periportal zones, a decrease in the activity of LH degradation, gross morphological changes in mitochondria, a decrease in the activity of surface LH degradation depend on the activity and duration of opioid dependence and are more pronounced with long-term (more than 6 years) highly progressive opioid dependence.

Histopathological findings of nonalcoholic fatty liver disease and nonalcoholic steatohepatitis.

Nonalcoholic fatty liver disease (NAFLD) is based on the concept of pathological morphology as well as clinical findings, and is broadly categorized into nonalcoholic fatty liver (NAFL) and nonalcoholic steatohepatitis (NASH). The differential diagnosis between NAFL and NASH is important because NASH has the potential to progress to cirrhosis and hepatocellular carcinoma. NAFL is simple hepatic steatosis without hepatocellular injury, while NASH is characterized by macrovesicular steatosis, inflammation, and ballooning hepatocytes with a predominantly centrilobular (zone 3) distribution. Liver biopsy is a useful test for diagnosing NAFLD, but it is invasive. Therefore, various noninvasive methods including diagnostic imaging have been developed in recent years. To verify their usefulness, it is necessary to clarify in detail how the pathological findings are reflected in the image findings as imaging and histopathological findings are closely related. We describe the main histological features of NAFLD, i.e., steatosis, inflammation, ballooning hepatocytes, Mallory-Denk bodies, and fibrosis, as well as the evolutional process to liver cirrhosis.

Dietary Iron Overload Differentially Modulates Chemically-Induced Liver Injury in Rats.

Hepatic iron overload is well known as an important risk factor for progression of liver diseases; however, it is unknown whether it can alter the susceptibility to drug-induced hepatotoxicity. Here we investigate the pathological roles of iron overload in two single-dose models of chemically-induced liver injury. Rats were fed a high-iron (Fe) or standard diet (Cont) for four weeks and were then administered with allyl alcohol (AA) or carbon tetrachloride (CCl4). Twenty-four hours after administration mild mononuclear cell infiltration was seen in the periportal/portal area (Zone 1) in Cont-AA group, whereas extensive hepatocellular necrosis was seen in Fe-AA group. Centrilobular (Zone 3) hepatocellular necrosis was prominent in Cont-CCl4 group, which was attenuated in Fe-CCl4 group. Hepatic lipid peroxidation and hepatocellular DNA damage increased in Fe-AA group compared with Cont-AA group. Hepatic caspase-3 cleavage increased in Cont-CCl4 group, which was suppressed in Fe-CCl4 group. Our results showed that dietary iron overload exacerbates AA-induced Zone-1 liver injury via enhanced oxidative stress while it attenuates CCl4-induced Zone-3 liver injury, partly via the suppression of apoptosis pathway. This study suggested that susceptibility to drugs or chemical compounds can be differentially altered in iron-overloaded livers.

Canine copper-associated hepatitis: A retrospective study of 17 clinical cases.

Background:Copper-associated hepatitis (CAH) is a well-documented chronic hepatic disease in dogs. In some breeds, the disease results from an inherited defect in copper metabolism. In others, it is unclear whether its acummulation is a primary or secondary condition. Reports of copper accumulation in dog breeds that are not genetically predisposed are increasing.Aim:To describe the epidemiology, clinical and laboratory findings, liver biopsy techniques, and treatment response in dogs with CAH.Methods:A retrospective study was performed, drawing upon medical records from CAH dogs at a Veterinary Referral Hospital in Paris, France. The diagnosis of CAH had been confirmed in these patients by positive rhodanine staining of hepatic tissue obtained through biopsy. Medical records were mined for the following data: age at presentation, sex, breed, chief presenting complaints, abdominal ultrasound (US) findings, and rhodanine staining pattern.Results:A total of 17 dogs were included in the study. Median age at presentation was 8-year old (4–11). No sex predisposition was found. Terriers (4/17) and German Shepherd Dogs (GSD, 3/17) were overrepresented. American Staffordshire Terriers and Beauceron had not previously appeared in case reports on CAH; two of each breed were identified in this study. Clinical signs of affected dogs were non-specific. An incidental identification of increased liver-enzymes was observed in 5/17 dogs. A heterogeneous, mottled liver was frequently described (5/17) on abdominal US. Liver biopsies were performed by US-guided percutaneous approach in 10/17 dogs, laparoscopy and laparotomy in 6/17 and 1/17, respectively. The rhodanine staining pattern was centrilobular (zone 3) in 8/17 dogs and periportal (zone 1) in 3/17 dogs. The pattern was considered multifocal in 6/17 dogs.Conclusion:Increased liver enzymes may be the only clinical finding in dogs with copper-associated hepatitis, reflecting the silent progression of this disease. Centrilobular pattern of rhodanine staining was observed in the majority of cases suggesting the primary condition of the disease. Results of this study are consistent with the current literature, which reports that terriers and GSD are predisposed to CAH. This is the first description of CAH in Beauceron and American Staffordshire Terrier dogs.

The ultrasonographic and pathomorphological markers of liver steatosis in patients with non-alcoholic fatty liver disease depending on the presence of comorbid chronic kidney disease (chronic pyelonephritis) and obesity

Objective — to establish the peculiarities of ultrasonographic and pathomorphological changes of liver parenchyma in patients with non-alcoholic liver steatosis (NAS) and non-alcoholic steatohepatitis (NASH) depending on the presence of comorbid chronic kidney disease (CKD) and obesity Materials and methods. The examinations involved 384 patients with non-alcoholic fatty liver disease (NAFLD), from them 84 subjects with I degree obesity (1 group) were divided into 2 subgroups: 32 patients with NAS and 52 patients with NASH. The second group consisted of 270 patients with NAFLD with comorbid I degree obesity and CKD І—ІІІ stage and included 110 patients with NAS and 160 patients with NASH. The control group consisted of 90 patients with CKD of the І—ІІІ stage with normal body weight (group 3). The mean age of patients was (45.8 ± 3.81) years. Results and discussion. The study demonstrated that the comorbidity of obesity and NASH with CKD increases the degree of steatosis (the area of hepatocytes affected by fatty liver dystrophy), the degree of ballooning degeneration, hyaline-drop dystrophy; the dystrophic changes in the centrilobular zone (3) deepen in the middle of the lobe with the development of panacinar injury and the involvement of zone 1. The course of NASH with CKD is characterized by a significantly higher incidence and intensity of clinical syndromes and the degree of biochemical and histological activity, which leads to the progression of fibrotic changes in the liver F2—F3. Conclusions. The NASH and CKD comorbidity is characterized by a higher degree of liver steatosis (LSD) (hepatorenal index 1.3 times higher than in the group of patients with NASH, p < 0.05) and the higher diagnostic threshold of values of the hepatotoxic index, which in strong interdependence correlates with the degree of steatosis of the liver, determined by Steato-test (r = 0.87; p < 0.001). In patients with NAS and NASH without comorbid pathology, a significant degree of LSD was found, with NASH it exceeds the reference values by 3.3 times (p < 0.05) and with NAS — by 1.8 times (p < 0.05). In comorbidity with CKD in patients with NAS, LSD was 3.5 times higher (p < 0.05) and in patients with NASH — 4.6 times higher (p < 0.05) than the reference values (p < 0.05).

Protective effect of tetrahydrobiopterin on hepatic and renal damage after acute cadmium exposure in male rats

ABSTRACTCadmium (Cd) has been recognized as one of the most important environmental and industrial pollutants. This study investigated the impact of acute exposure to Cd on oxidative stress and the inflammatory marker interleukin-6 (IL-6) in the plasma of rats and the histological picture of liver and kidney, as well as to examine the potential protective effect of tetrahydrobiopterin (BH4). Methods: Rats were divided into control group, Cd group that received a single intraperitoneal (i.p.) dose of 4 mg/kg b.w. of CdCl2 and BH4+ Cd group that received a single dose of BH4 (20 mg/kg, i.p.) and subsequently exposed to a single dose of Cd 24 h after the BH4 treatment. Results: Cd increased the plasma levels of hepatic enzymes (ALT and AST), urea, creatinine, malondialdehyde (MDA), and IL-6 and decreased the superoxide dismutase (SOD) activity. Also, it induced histopathological alterations in the liver with severe degeneration, especially in centrilobular zones. Renal tubular epithelium showed vacuolated cytoplasm and dense nuclei. VEGF expression was mild. Ultrastuctural changes were seen in some renal tubules. The nuclei appeared distorted with electron dense chromatin. Mitochondria with destructed cristae were observed. BH4 pretreatment had protective effects, since it significantly reduced the levels of IL-6 and ameliorated the alteration in oxidative status biomarkers induced by Cd. Improvement of histopathological alterations was observed in Cd-groups. The nuclei were vesicular euchromatic, intact mitochondria and normal appearance of the filtration membrane. Moderate expression of VEGF was noted. Conclusion: This study has provided clear evidence for the protective efficacy of BH4 against experimental Cd toxicity.

Centrilobular ductular reaction correlates with fibrosis stage and fibrosis progression in non-alcoholic steatohepatitis

There is increasing interest in the role of ductular reaction as part of the pathogenesis and characteristic histology of non-alcoholic steatohepatitis. However, earlier studies did not separately assess the contribution of periportal and centrilobular zone ductular reaction over the spectrum of non-alcoholic steatohepatitis, and their clinical significance remains unclear. We herein analyzed the character of ductular reaction in each hepatic zone in non-alcoholic steatohepatitis biopsies and for the first time evaluated the prognostic value of ductular reaction in baseline biopsies as a predictor of progression of fibrosis in subsequent biopsies. A total of 90 non-alcoholic steatohepatitis liver biopsies were included in the cohort. The relationships among ductular reaction, grade, stage, and other common histopathologic findings in non-alcoholic steatohepatitis were analyzed in a cross-sectional manner. Among these patients, a total of 47 patients underwent sequential liver biopsies in the absence of effective treatment. The frequency of ductular reaction and the other histopathologic parameters in the initial biopsies were analyzed as predictors of progression of fibrosis in the second biopsies in a longitudinal analysis. Centrilobular ductular reaction was identified in 90% of patients and 38% of centrilobular zones. The prevalence of centrilobular ductular reaction increased as non-alcoholic steatohepatitis grade increased (P=0.0002) and also as stage of fibrosis increased (P<0.0001) in the cross-sectional study. In the longitudinal study, the frequency of centrilobular ductular reaction in the initial biopsies was significantly higher in the group of progressors and correlated with the rate of fibrosis progression (P=0.02). Centrilobular ductular reaction is common in non-alcoholic steatohepatitis and its presence correlates significantly with increasing necroinflammatory activity and fibrosis stage. Development of centrilobular ductular reaction appears to predict progression of fibrosis in subsequent biopsies.

Endocrine disrupting chemicals, 4-nonylphenol, bisphenol A and butyl benzyl phthalate, impair metabolism of estradiol in male and female rats as assessed by levels of 15α-hydroxyestrogens and catechol estrogens in urine.

Bisphenol A (BPA), 4-nonylphenol (NP) and butyl benzyl phthalate (BBP), termed endocrine-disrupting chemicals, are known to mimic estrogen activity. The effects of these chemicals on 17β-estradiol (E2 ) metabolism in vivo in rats were examined. Male and female rats were given NP (250mg kg-1 day-1 ), BPA (250μg kg-1 day-1 ) or BBP (500mg kg-1 day-1 ) by gavage for 14days, followed by a single intraperitoneal injection of E2 (5mg kg-1 ) on the final day. The urinary excretion over 72hours of 2-hydroxyestrone 1-N-acetylcysteine thioether, 2-hydroxyestrone 4-N-acetylcysteine thioether, 4-hydroxyestrone 2-N-acetylcysteine thioether, 2-hydroxy-17β-estradiol (2-OHE2 ), 2-hydroxyestrone (2-OHE1 ), 4-hydroxy-17β-estradiol, 4-hydroxyestrone, 15α-hydroxyestriol (E4 ), 15α-hydroxy-17β-estradiol and 15α-hydroxyestrone was measured. Increases in urinary excretion of 2-OHE1 and decreases in E4 were observed in males treated with NP or BBP. Decreases in urinary excretion of 2-OHE2 and E4 were observed in males treated with BPA. Decreases in urinary excretion of 2-OHE1 and 2-OHE2 were observed in females treated with BBP. Normalized liver and weights were increased in both sexes treated with NP or BBP. Histologic observations revealed marked changes in the distal tubules and collecting ducts in the kidneys of rats exposed to NP and BBP, and hypertrophy in the hepatocytes of the centrilobular zone of the liver. No BPA-related effects on organ weight and on liver or kidney histopathology were found. These results suggest that the 14day oral dosing of NP and BBP disrupted E2 metabolism, resulting from marked morphological and functional alterations in the liver and kidneys. In addition, BPA could induce metabolic and endocrine disruption.

Identification of miR-31-5p, miR-141-3p, miR-200c-3p, and GLT1 as human liver aging markers sensitive to donor-recipient age-mismatch in transplants.

SummaryTo understand why livers from aged donors are successfully used for transplants, we looked for markers of liver aging in 71 biopsies from donors aged 12–92 years before transplants and in 11 biopsies after transplants with high donor–recipient age‐mismatch. We also assessed liver function in 36 age‐mismatched recipients. The major findings were the following: (i) miR‐31‐5p, miR‐141‐3p, and miR‐200c‐3p increased with age, as assessed by microRNAs (miRs) and mRNA transcript profiling in 12 biopsies and results were validated by RT–qPCR in a total of 58 biopsies; (ii) telomere length measured by qPCR in 45 samples showed a significant age‐dependent shortage; (iii) a bioinformatic approach combining transcriptome and miRs data identified putative miRs targets, the most informative being GLT1, a glutamate transporter expressed in hepatocytes. GLT1 was demonstrated by luciferase assay to be a target of miR‐31‐5p and miR‐200c‐3p, and both its mRNA (RT–qPCR) and protein (immunohistochemistry) significantly decreased with age in liver biopsies and in hepatic centrilobular zone, respectively; (iv) miR‐31‐5p, miR‐141‐3p and miR‐200c‐3p expression was significantly affected by recipient age (older environment) as assessed in eleven cases of donor–recipient extreme age‐mismatch; (v) the analysis of recipients plasma by N‐glycans profiling, capable of assessing liver functions and biological age, showed that liver function recovered after transplants, independently of age‐mismatch, and recipients apparently ‘rejuvenated’ according to their glycomic age. In conclusion, we identified new markers of aging in human liver, their relevance in donor–recipient age‐mismatches in transplantation, and offered positive evidence for the use of organs from old donors.

Oxaliplatin-based chemotherapy induces extravasated platelet aggregation in the liver.

Oxaliplatin-based chemotherapy plays a central role in the treatment of patients with colorectal liver metastasis (CRLM). This treatment, however, has been associated with hepatic sinusoidal obstruction syndrome (SOS), a clinically important adverse effect characterized by a bluish hue of the liver, splenomegaly and thrombocytopenia, resulting in liver dysfunction. The significant association between the sinusoidal endothelium and platelets has suggested that oxaliplatin-based chemotherapy affects platelets in the liver. This study compared platelet counts in patients who did and did not receive oxaliplatin-based neoadjuvant chemotherapy (NAC). The peripheral blood platelet count was significantly lower in the NAC group (n=17) compared to that in the non-NAC, or control group (n=15) (P<0.05). The spleen index was also higher in the NAC group, although the difference was not significant. However, the spleens of the patients in the NAC group were significantly enlarged following treatment (P<0.01). Immunostaining for the platelet surface marker CD42b (glycoprotein Ib), revealed more platelets in the liver in the NAC compared to the control group, particularly in the centrilobular zone III, adjacent to the hepatic central vein and in contact with hepatocytes (P<0.01). The platelets present in the spaces of Disse, referred to as extravasated platelet aggregation (EPA), secrete a number of growth factors, including transforming growth factor-β, vascular endothelial growth factor-A, plasminogen activator inhibitor-1 and thromboxane A2. In conclusion, EPA may play an important role in the development of hepatic SOS. Moreover, antiplatelet drugs may prevent the onset of SOS and hepatic injury in patients treated with oxaliplatin-based chemotherapy for CRLM.

Death due to acute fatty liver in pregnancy

Introduction: Acute fatty liver of pregnancy (AFLP) results due to disordered metabolism of fatty acids by mitochondria in the mother, caused by deficiency ofdehydrogenase enzyme. This is a rare instance of a foetus causing metabolic disease in the mother. The ‘working diagnosis’ of the clinicians was HELLP syndrome. The exact cause of death was made clear only after the postmortem examination. Case report : 24 year old primi with 34 weeks of gestation admitted for blood pressure monitoring. She developed jaundice, high blood pressure with proteinurea, marginally elevated liver enzymes, low platelets which warranted termination of pregnancy. Despite the termination of pregnancy, she developed renal failure and hepatic encephalopathy and died on the eleventh day after delivery. Post mortem histopathology of liver revealed that the architecture was not altered and micro-vesicular fat infiltrations in centrilobular and mid zones. Hepatocytes were swollen but inflammation was not evident. Other tissues did not show any fat infiltration. HELLP syndrome was excluded due to absence of macro-vesicular fat infiltration in modest quantities throughout the liver lobules. There were no lobular disarray with hepatocyte dropout, reticulin collapse and portal tract inflammation to suggest viral hepatitis. There were no evidence of drug induced, toxic or viral induced micro-vesicular steatosis. Reye’s syndrome is characterized by uniform micro-vesicular fatty change in the liver and other tissues such as skeletal muscles, kidney and heart. Conclusion: Having excluded all other possibilities, the cause of death was ascertained as AFLP and this emphasizes the value of correct histopathological diagnosis . Medico-Legal Journal of Sri Lanka 2013; 1(2) : 42-45