The effects of 48 weeks of rosiglitazone on hepatocyte mitochondria in human nonalcoholic steatohepatitis

Rosiglitazone, a thiazolidinedione peroxisome proliferator-activated receptor gamma ligand, reduces disease activity in nonalcoholic steatohepatitis (NASH), a disease associated with hepatocyte mitochondrial crystalline inclusions that are not seen in animal models of NASH. In human and animal studies of adipose tissue, thiazolidinediones may induce mitochondrial biogenesis and associated morphological changes. To determine if rosiglitazone alters the hepatocyte mitochondrial morphology in human NASH, we prospectively and systematically examined liver biopsies from human subjects with NASH before and after 48 weeks of rosiglitazone by transmission electron microscopy. Twenty patients (body mass index = 34 +/- 7) were studied. Four coded sections from each of 20 pretherapy biopsies and each of 20 posttherapy biopsies were examined by transmission electron microscopy. The total hepatocyte mitochondria and crystal-containing mitochondria were counted, and semiquantitative scoring was performed for macrosteatosis, microsteatosis, dilated endoplasmic reticulum, apoptosis, Mallory bodies, and hepatocyte enlargement. The total mitochondria count was unchanged after therapy, but there was a significant increase in crystal-containing mitochondria from 4.0% (95% confidence interval = 1.8-8.8) to 7.2% (95% confidence interval = 3.9-12.6; odds ratio = 1.80; P = 0.04) after the treatment with rosiglitazone. Macrosteatosis (P < 0.001) and Mallory bodies (P = 0.05) significantly decreased, but no change was evident in microsteatosis, cellular enlargement, dilated endoplasmic reticulum, or apoptosis. CONCLUSION: Rosiglitazone therapy of NASH is associated with increased crystalline inclusions in hepatocyte mitochondria. Whether these are adaptive or pathological remains unknown, and further studies are warranted to assess hepatic mitochondrial function during thiazolidinedione therapy for NASH.     

Pediatric non-alcoholic steatohepatitis:The first report on the ultrastructure of hepatocyte mitochondria

AIM:To investigate the ultrastructure of abnormal hepatocyte mitochondria,including their cellular and hepatic zonal distribution,in bioptates in pediatric nonalcoholic steatohepatitis(NASH).METHODS:Ultrastructural investigations were conducted on biopsy liver specimens obtained from 10children(6 boys and 4 girls)aged 2-14 years with previously clinicopathologically diagnosed NASH.The disease was diagnosed if liver biopsy revealed steatosis,inflammation,ballooned hepatocytes,Mallory hyaline,or focal necrosis,varying degrees of fibrosis in the absence of clinical,serological,or histological findings of infectious liver diseases,autoimmune hepatitis,metabolic liver diseases,or celiac disease.For ultrastructural analysis,fresh small liver blocks(1 mm3 volume)were fixed in a solution containing 2%paraformaldehyde and 2.5%glutaraldehyde in 0.1 mol/L cacodylate buffer.The specimens were postfixed in osmium tetroxide,subsequently dehydrated through a graded series of ethanols and propylene oxide,and embedded in Epon812.The material was sectioned on a Reichert ultramicrotome to obtain semithin sections,which were stained with methylene blue in sodium borate.Ultrathin sections were contrasted with uranyl acetate and lead citrate,and examined using an Opton EM 900 transmission electron microscope.RESULTS:Ultrastructural analysis of bioptates obtained from children with non-alcoholic steatohepatitis revealed characteristic repetitive mitochondrial abnormalities within hepatocytes;mainly mitochondrial polymorphisms such as megamitochondria,loss of mitochondrial cristae,and the presence of linear crystalline inclusions within the mitochondrial matrix of an increased electron density.The crystalline inclusions were particularly evident within megamitochondria(MMC),which seemed to be distributed randomly both within the hepatic parenchymal cell and the zones of hepatic lobule,without special variations in abundance.The inclusions appeared as bundles viewed longitudinally,or as an evenly spaced matrix in cross section,and frequently caused mitochondrial deformation.The average diameter of these linear structures was 10 nm and the average space between them 20 nm.Sometimes enlarged intramitochondrial granules were seen in their vicinity.Foamy cytoplasm of hepatocytes was found,resulting from the proliferation of smooth endoplasmic reticulum and glycogen accumulation.The perivascular space of Disse was frequently dilated,and contained transitional hepatic stellate cells,as well as mature and/or newly forming collagen fiber bundles.CONCLUSION:Marked ultrastructural abnormalities observed in hepatocyte mitochondria,especially their polymorphism in the form of MMC and loss of mitochondrial cristae,accompanied by foamy cytoplasm,clearly indicate a major role of these organelles in the morphogenesis of pediatric NASH.Our findings seem to prove the high effectiveness of electron microscopy in the diagnosis of the disease.     

Electron microscopic findings in non‐alcoholic fatty liver disease: Is there a difference between hepatosteatosis and steatohepatitis?

Background and Aims: Non‐alcoholic fatty liver disease has long been accepted as benign; however, recent evidence suggests that the disease may progress to cirrhosis and hepatocellular carcinoma, although the natural course of the disease is still unclear. This study was designed to comparatively evaluate electron microscopic features of non‐alcoholic fatty liver (NAFL) and non‐alcoholic steatohepatitis (NASH). Methods: Quantitative and semi‐quantitative ultrastructural evaluations were performed on liver biopsies from 23 patients, 10 with NAFL and 13 with NASH. Results: No statistically significant difference was noted between NAFL and NASH patients in ultrastructural features of hepatocytes including megamitochondria, intramitochondrial crystalline inclusions, mitochondrial matrix granules, foamy cytoplasmic appearance, electron‐lucent and glycogen‐containing nuclear regions, lipofuscin granules, or an increased frequency of vesicles containing electron‐dense material in peribiliary Golgi zone; however, the mitochondrial diameter was significantly higher in the NASH patients. Intercellular distance and microvilli between hepatocytes, collagen and electron‐dense material accumulation in the space of Disse, electron‐dense material accumulation and microvillus density in bile canaliculi did not differ significantly between the groups. Conclusions: Our data show that, although NAFL and NASH can be distinguished by their distinct light microscopic features, ultrastructural characteristics are similar, which suggests that NAFL may also have the potential to progress to fibrosis and cirrhosis like NASH.     

Mitochondrial creatine kinase: a major constituent of pathological inclusions seen in mitochondrial myopathies.

Overaccumulation of abnormally organized mitochondria in so-called "ragged-red" skeletal muscle fibers is a morphological hallmark of mitochondrial myopathies, in particular of mitochondrial encephalomyopathies. Characteristic for the abnormal mitochondria is the occurrence of highly ordered crystalline inclusions. Immuno-electron microscopy revealed that these inclusions react heavily with specific antibodies against mitochondrial creatine kinase (Mi-CK). Image processing of selected crystalline inclusions, sectioned along the crystallographic b, c planes, resulted in an averaged picture displaying an arrangement of regular, square-shaped particles with a central cavity. The overall appearance, dimensions, and symmetry of these building blocks are very reminiscent of single isolated Mi-CK octamers. Taking these findings together, it is concluded that Mi-CK octamers indeed represent the major, if not the only, component of these mitochondrial inclusions.     

Proteomic analysis of liver mitochondria from rats with nonalcoholic steatohepatitis

AIM:To explore mitochondrial dysfunction in nonalcoholic steatohepatitis(NASH)by analyzing the proteome of liver mitochondria from a NASH model.METHODS:The NASH rat model was established by feeding rats a fat-rich diet for 24 wk and was confirmed using hematoxylin and eosin staining of liver tissue and by changes in the levels of serum alanine transaminase,aspartate aminotransferase,triglyceride,total cholesterol and other markers.Liver mitochondria from each group were isolated using differential centrifugation.The mitochondrial samples were lyzed,purified and further analyzed using two-dimensional electrophoresis combined with matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry.Bioinformatic analyses of assigned gene ontology and biological pathway was used to study functional enrichments in the abundant proteomic data.RESULTS:Eight up-regulated and sixteen down-regulated proteins were identified that showed greater than1.5-fold differences between the controls and the NASH group.These dysregulated proteins were predicted to be involved in different metabolic processes including fatty acidβ-oxidation processes,lipid metabolic processes,cell-cycle arrest,cell polarity maintenance,and adenosine triphosphate/sex hormone metabolic processes.Novel proteins that may be involved in NASH pathogenesis including the trifunctional enzyme Hadha,thyroxine,prohibitin,aldehyde dehydrogenase ALDH1L2,UDP-glucuronosyltransferase 2B31,and carbamoyl-phosphate synthase were identified using bioinformatics tools.The decreased expression of Hadha in NASH liver was verified by Western blotting,which was used as a complementary technique to confirm the proteomic results.CONCLUSION:This novel report on the liver mitochondrial proteome of a NASH model may provide a reservoir of information on the pathogenesis and treatment of NASH.     

Mitochondrial hepatopathies: advances in genetics and pathogenesis

Hepatic involvement is a common feature in childhood mitochondrial hepatopathies, particularly in the neonatal period. Respiratory chain disorders may present as neonatal acute liver failure, hepatic steatohepatitis, cholestasis, or cirrhosis with chronic liver failure of insidious onset. In recent years, specific molecular defects (mutations in nuclear genes such as SCO1, BCS1L, POLG, DGUOK, and MPV17 and the deletion or rearrangement of mitochondrial DNA) have been identified, with the promise of genetic and prenatal diagnosis. The current treatment of mitochondrial hepatopathies is largely ineffective, and the prognosis is generally poor. The role of liver transplantation in patients with liver failure remains poorly defined because of the systemic nature of the disease, which does not respond to transplantation. Prospective, longitudinal, multicentered studies will be needed to address the gaps in our knowledge in these rare liver diseases.     

Changes in human hepatic metabolism in steatosis and cirrhosis

AIM To understand the underlying metabolic changes in human liver disease we have applied nuclear magnetic resonance(NMR) metabolomics analysis to human liver tissue.METHODS We have carried out pilot study using 1H-NMR to derive metabolomic signatures from human liver from patients with steatosis, nonalcoholic steatohepatitis(NASH) or alcohol-related liver damage(ARLD) to identify species that can predict outcome and discriminate between alcohol and metabolic-induced liver injuries. RESULTS Changes in branched chain amino acid homeostasis, tricarboxylic acid cycle and purine biosynthesis intermediates along with betaine were associated with the development of cirrhosis in both ARLD and nonalcoholic fatty liver disease. Species such as propylene glycol and as yet unidentified moieties that allowed discrimination between NASH and ARLD samples were also detected using our approach.CONCLUSION Our high throughput, non-destructive technique for multiple analyte quantification in human liver specimens has potential for identification of biomarkers with prognostic and diagnostic significance.     

Nonalcoholic steatohepatitis: association of insulin resistance and mitochondrial abnormalities

BACKGROUND AND AIMS: The pathogenesis of nonalcoholic steatohepatitis (NASH) is unknown. We tested the hypothesis that NASH is associated with 2 defects: (1) peripheral insulin resistance, which increases lipolysis, delivery of free fatty acids (FFA) to the liver, and hepatic fatty acid beta oxidation, thereby creating oxidative stress; and (2) an abnormality within the hepatocytes that might render them more susceptible to injury from oxidative stress. METHODS: The hypothesis was tested by evaluation of (1) insulin resistance by a 2-step hyperinsulinemic (10 and 40 mU. m(-2). min(-1)) euglycemic clamp; (2) insulin effects on lipolysis by enrichment of [U-(13)C]glycerol; (3) frequency and severity of structural defects in hepatocyte mitochondria in vivo; (4) fatty acid beta oxidation from serum [beta-OH butyrate], release of water-soluble radioactivity from (3)H-palmitate by cultured fibroblasts and urinary dicarboxylic acid excretion; and (5) hepatic lipid peroxidation by immunohistochemical staining for 3-nitrotyrosine (3-NT). Subjects with NASH (n = 6-10 for different studies) were compared with those with fatty liver (n = 6) or normal controls (n = 6). RESULTS: NASH and fatty liver were both associated with insulin resistance, with mean glucose infusion rates (normal/fatty liver/NASH) of step 1, 4.5/1.6/0.9; step 2, 9.5/7.7/4.5 (P < 0.03 for both steps). Although baseline rates of glycerol appearance were higher in those with NASH than in those with fatty liver (means, 14.6 vs. 21.6 micromol. kg(-1). min(-1); P < 0.05), neither group significantly suppressed glycerol appearance at insulin infusion rates of 10 mU. m(-2). min(-1). NASH was associated with loss of mitochondrial cristae and paracrystalline inclusions in 9 of 10 subjects, compared with 0 of 6 subjects with fatty liver. However, no evidence of a generalized defect in fatty acid beta oxidation was noted in any group. Also, mean [beta-OH butyrate] was highest in those with NASH (means, 90 vs. 110 vs. 160 micromol/L; P < 0.04). Increased staining for 3-NT was present in fatty liver, and even greater staining was seen in NASH. CONCLUSIONS: These data indicate that peripheral insulin resistance, increased fatty acid beta oxidation, and hepatic oxidative stress are present in both fatty liver and NASH, but NASH alone is associated with mitochondrial structural defects.     

Hepatocyte apoptosis and fas expression are prominent features of human nonalcoholic steatohepatitis

BACKGROUND & AIMS: The pathogenesis of nonalcoholic steatohepatitis (NASH) remains poorly understood. Although apoptosis is a common mechanism of liver injury, the extent and clinical significance of apoptosis in NASH has not been examined. Thus, the aims of this study were to quantify hepatocyte apoptosis in NASH, correlate it with disease severity, and identify possible mechanisms of apoptosis induction. METHODS: Hepatocyte apoptosis was assessed in NASH, simple steatosis, alcoholic hepatitis, and controls without liver disease using the TUNEL assay and immunohistochemistry for activated caspases 3 and 7. Liver specimens were also graded according to the magnitude of inflammation and fibrosis. RESULTS: TUNEL-positive cells were significantly increased in liver biopsy specimens from patients with NASH compared with simple steatosis and controls. Unexpectedly, TUNEL-positive cells were also greater in NASH vs. alcoholic hepatitis. Immunohistochemistry demonstrated active caspases 3 and 7 in NASH specimens, confirming the occurrence of apoptosis in this disease. A positive correlation was observed between hepatocyte apoptosis and hepatic fibrosis and inflammatory activity, respectively. The Fas receptor was strongly expressed in hepatocytes in liver specimens from NASH patients as compared with controls. CONCLUSIONS: Hepatocyte apoptosis is significantly increased in patients with NASH and correlates with disease severity, suggesting that antiapoptotic therapy may be useful in this syndrome.     

Mitochondrial antiviral signaling protein defect links impaired antiviral response and liver injury in steatohepatitis in mice

Mitochondrial dysfunction is a pathogenic feature of nonalcoholic steatohepatitis (NASH). NASH complicates hepatotropic viral disease. The mitochondrial antiviral signaling protein (MAVS) is the adapter of helicase receptors involved in sensing double-stranded RNA (dsRNA). We hypothesized that impaired MAVS function may contribute to insufficient antiviral response and liver damage in steatohepatitis. We identified reduced MAVS protein levels and increased MAVS association with the proteasome subunit alpha type 7 (PSMA7) in livers from mice given a methionine-choline-deficient (MCD) diet. Decreased association of MAVS with mitochondria and increased cytosolic cytochrome c indicated mitochondrial damage in steatohepatitis. In vivo administration of the synthetic dsRNA polyinosinic:polycytidylic acid [poly(I:C)], but not lipopolysaccharide or cytidine-phosphate-guanosine-rich DNA, resulted in impaired induction of type I interferons (IFNs) and proinflammatory cytokines in steatohepatitis. Consistent with a defect in helicase receptor-induced signaling, there was loss of poly(I:C)-induced translocation of MAVS to the cytosol and decreased IFN regulatory factor 3 phosphorylation. Caspases 1 and 8, both of which cleave MAVS, were increased in MCD diet-fed mice. At baseline, steatohepatitis was associated with increased serum alanine aminotransferase (ALT), apoptosis and caspase 3 activation compared with controls. In contrast to apoptosis in controls, necrosis was induced by poly(I:C) stimulation in steatohepatitis. Hepatocyte necrosis was indicated by elevated serum high-mobility group box protein-1 and ALT and was correlated with increased expression of receptor-interacting protein 3 (RIP3), a master regulator of necrosis. Increased expression of MAVS, PSMA7, and RIP3 messenger RNA was also present in human NASH livers. CONCLUSION: Our novel findings suggest that mitochondrial damage in steatohepatitis extends to MAVS, an adapter of helicase receptors, resulting in inefficient type I IFN and inflammatory cytokine response but increased hepatocyte necrosis and RIP3 induction in response to a dsRNA viral challenge. These mechanisms may contribute to progressive liver damage and impaired viral clearance in NASH.     

Hepatic gene expression in patients with obesity-related non-alcoholic steatohepatitis.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is among the most common causes of chronic liver disease. NAFLD includes a spectrum of clinicopathologic syndromes that includes non-alcoholic steatohepatitis (NASH) that has potential for progression. The pathogenesis of NASH is poorly characterized. AIM: This study was designed to identify differences in hepatic gene expression in patients with NASH and to relate such differences to their clinical characteristics. DESIGN: Consecutive patients undergoing bariatric surgery were prospectively recruited. Extensive clinical data and two liver biopsy specimens were obtained at the time of enrollment. A single hepatopathologist reviewed and classified the liver biopsies. Patients with excessive alcohol use and other causes of liver disease were excluded. A group of 29 NASH patients, 12 with steatosis alone, seven obese controls and six non-obese controls were selected for further investigation. Customized cDNA microarrays containing 5220 relevant genes were designed specifically for this study. Microarray experiments were run in triplicate for each sample and a selected group of genes were confirmed using real-time PCR. OUTCOME MEASURE: Differential hepatic gene expressions in patients with NASH as compared with controls. Results: Thirty-four genes with significant differential expression were identified in patients with NASH when compared with non-obese controls. Moreover, 19 of these genes showed no significant expression differences in obese vs. non-obese controls, suggesting a stronger association of these genes to NASH. CONCLUSIONS: Several differentially expressed genes in patients with NASH are related to lipid metabolism and extracellular matrix remodeling. Additionally, genes related to liver regeneration, apoptosis, and the detoxification process were differentially expressed. These findings may help clarify the molecular pathogenesis of NASH and identify potential targets for therapeutic intervention.     

Abnormal electron microscopic findings of nonalcoholic steatohepatitis and related factors]

BACKGROUND/AIMS: In spite of increasing interests about nonalcoholic steatohepatitis (NASH), there are few reports about the ultrastructure of hepatocyte in this disease. The aim of this study was to clarify abnormal electron microscopic (EM) findings and related factors in NASH. METHODS: Total of fourteen patients who underwent liver biopsy due to steatohepatitis were included. Precise personal history was taken and variable blood tests such as liver function test, lipid profile, and serum iron study were done. Pathologic examination with light and electron microscopy was done by single pathologist. RESULTS: Eleven men and three women were included and mean age was 33.7+/-12.8 years. Nine patients drinking less than 40 g/week was grouped as "NASH group" and other 5 patients drinking more than 40 g/week and body mass index less than 25 was grouped as "ASH (Alcoholic Steatohepatitis) group". Polymorphism of mitochondria such as megamitochondria or loss of cristae was major abnormal EM findings and was more common in "NASH group" than "ASH group" (p=0.027). There was no significant clinical or pathological factors related with the presence of these abnormal EM findings. CONCLUSIONS: Polymorphism of mitochondria is major abnormal EM finding of steatohepatitis and is more common in NASH than ASH. And there is no significant clinical or pathological factors which could predict the presence of these abnormal EM findings.     

Increased expression of cytochrome P450 2E1 in nonalcoholic fatty liver disease: mechanisms and pathophysiological role

Due to the worldwide surge in obesity and type 2 diabetes, the increased incidence of nonalcoholic fatty liver disease (NAFLD) is a major concern for the public health. Indeed, NAFLD encompasses a large spectrum of conditions ranging from fatty liver to nonalcoholic steatohepatitis (NASH), which can progress to cirrhosis in some patients. A better understanding of the mechanisms involved in fatty liver and its progression into NASH is important in order to develop efficient drugs able to alleviate these liver diseases. Although numerous investigations pointed to reactive oxygen species (ROS) as key players in the progression of fatty liver to NASH, their exact source is still uncertain. Besides the mitochondrial respiratory chain, cytochrome P450 2E1 (CYP2E1) has recently emerged as another potentially important cause of ROS overproduction. Indeed, higher hepatic CYP2E1 expression and activity have been frequently observed in the context of obesity and NAFLD. It is currently unknown why CYP2E1 is enhanced in these dysmetabolic diseases, although increased hepatic levels of fatty acids and insulin resistance might play a role. Nonetheless, higher hepatic CYP2E1 could play a significant role in the pathophysiology of NASH by inducing lipid peroxidation and oxidative damage of key cellular components. Moreover, CYP2E1-mediated overproduction of ROS could promote hepatic insulin resistance, which can further aggravate fatty liver. Since a significant amount of CYP2E1 can be located within liver mitochondria, higher levels of CYP2E1 in NAFLD could also have detrimental effects on mitochondrial function. Finally, increased CYP2E1 activity during NAFLD could enhance the susceptibility of some patients to the hepatotoxicity of different xenobiotics through the CYP2E1-mediated generation of harmful reactive metabolites.     

Cimetidine hepatitis

A 62-year-old man developed asymptomatic anicteric hepatitis during cimetidine therapy for duodenal ulcer disease. Liver biopsy revealed zonal (centrolobular) necrosis, bile stasis, and mononuclear cell infiltration. Electron microscopy featured mitochondrial hyperplasia, giant mitochondria, and liver cell bile inclusions. The diagnosis of cimetidine-related hepatitis was confirmed by re-exposure to cimetidine, which prompted a rapid rise in serum transaminases while under observation in the hospital. Cimetidine hepatitis is rarely reported, and routine monitoring of the patient's liver tests during therapy does not seem cost-effective as a general practice.     

Usefulness of Technetium-99 m-2-methoxy-isobutyl-isonitrile liver scintigraphy for evaluating disease activity of non-alcoholic fatty liver disease

Aim: Non‐alcoholic steatohepatitis (NASH) is a progressive form of non‐alcoholic fatty liver disease (NAFLD). Therefore, it is important to evaluate disease activity and distinguish NASH from simple steatosis in NAFLD. Technetium‐99 m‐2‐methoxy‐isobutyl‐isonitrile (^99mTc‐MIBI) is a lipophilic cation designed for myocardial perfusion scintigraphy in the diagnosis of ischemic heart diseases, and its retention reflects mitochondrial function. It was reported that hepatic mitochondrial abnormalities would be an important predictive factor for NASH disease progression. The aim of this study was to examine the clinical usefulness of ^99mTc‐MIBI liver scintigraphy for evaluating disease activity of NAFLD and distinguishing NASH from simple steatosis in patients with NAFLD. Methods: Twenty‐six patients with biopsy‐proven NAFLD were enrolled. Clinicolaboratory tests and ^99mTc‐MIBI liver scintigraphy were performed. To evaluate hepatic uptake, regions of interest were set at the liver and heart, and the uptake ratio of the liver to heart (liver/heart ratio) was calculated. Results: All patients with NAFLD were classified into three groups according to the NAFLD activity score: non‐NASH (simple steatosis) (n = 4), borderline NASH (n = 11), and NASH (n = 11). Liver/heart ratios were significantly lower in NASH than in simple steatosis (P < 0.05). Moreover, liver/heart ratios were significantly correlated with NAFLD activity scores among the patients (r = ?0.413, P < 0.05). Conclusions: The present study indicates that ^99mTc‐MIBI liver scintigraphy would be a useful non‐invasive functional imaging method with which to evaluate disease activity of NAFLD and distinguish NASH from simple steatosis.     

Different pathophysiological mechanisms of intramitochondrial iron accumulation in acquired and congenital sideroblastic anemia caused by mitochondrial DNA deletion

Sideroblastic anemias (SA) are characterized by iron accumulation in the mitochondria of erythroblasts. Although we have evidence of mitochondrial gene alterations in sporadic congenital cases, the origin of acquired forms [refractory anemia with ring sideroblasts (RARS)], is still largely unknown. Here, we report the analysis of respiratory chain function in a patient with a large mitochondrial deletion and in patients with RARS. A young boy with SA showed symptoms typical of a mitochondrial disease with metabolic acidosis, muscle weakness and cerebral involvement. His bone marrow DNA was analyzed for the presence of mitochondrial deletions. We found a new mitochondrial (mt)DNA deletion spanning 3,614 bp and including all the mt genes encoding complex IV, plus ATPase 6 and 8, and several transfer (t)RNAs. All tissues analyzed (liver, skeletal muscle, brain, pancreas) showed a heteroplasmic distribution of this mutant DNA. Bone marrow homogenates were obtained from five patients with RARS and from three patients with normal bone marrow and respiratory chain function assayed by spectrophotometric analysis. Cytochrome c oxidase (CCO) activity was greatly reduced in the patient's bone marrow. In contrast, CCO activity and global respiratory chain function were conserved in patients with RARS. We conclude that deficient CCO activity secondary to mtDNA deletions is related to intramitochondrial iron accumulation, as in our patient or in those with Pearson's syndrome, whereas other mechanisms, e.g. nuclear DNA mutations, have to be proposed to be involved in the acquired forms of SA.     

Familial Mediterranean fever: an association with non-alcoholic fatty liver disease

The purpose of this study is to characterize the chronic liver disease (CLD) that may be associated with familial Mediterranean fever (FMF). Twenty-seven patients (mean age, 48 ± 18 years; F/M, 16:11) with FMF who were referred for assessment of CLD were studied. Data regarding FMF and CLD were obtained from patient medical files. Liver biopsy was performed in 21 of 27 patients and deferred in six (cirrhotic coagulopathy in five and one who improved after colchicine dose reduction). Patients with FMF and non-alcoholic fatty liver disease (NAFLD) were compared to matched controls from a cohort of 150 patients with NAFLD per liver biopsy but without FMF. The mean Tel Hashomer severity score was 1.7 ± 0.9. The mean daily dose of colchicine was 1.4 ± 0.4 mg over a mean duration of 21 years ± 10. Seven of ten patients who underwent mutation analysis for FMF were homozygous for M694V. In 15 patients, there was evidence of NAFLD: five with “simple” steatosis, three with non-alcoholic steatohepatitis (NASH), and seven with NASH-cirrhosis. An additional five patients had “cryptogenic” cirrhosis, which in most patients represents the end result of unrecognized NASH, and one had normal liver tissue. Comparing FMF patients with NAFLD to matched controls with NAFLD did not reveal excess of metabolic syndrome in FMF patients. Of our FMF patients, 74% had evidence of NAFLD, 75% of which with severe manifestation. The extremely high proportion of NAFLD in our cohort of FMF patients without overt metabolic syndrome may indicate an unappreciated novel association between FMF and NAFLD.